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Test 2 Flashcards

1
Q

what is adsorption

A

adhesion of molecules to solid surface (no penetration)

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2
Q

what affects adsorption the most

A

hydrophobicity and surface charge

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3
Q

protein adsorption increases with increasing __

A

hydrophobicity

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4
Q

besides hydrophobicity and surface charge, __ can also promote protein adsorption becuause __

A

surface roughness can promote adsorption because proteins will become physically trapped in the valleys of the surface

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5
Q

what are the chemical and physical properties governing protein adsorption

A 
chemcial
     Hydrophobicity
     Surface Charge
Physical
     Surface Roughness
     Steric hinderances
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6
Q

rougher surfaces will have a ___ protein adsorption

A

more protein adsorption

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7
Q

besides making a surface smoother how do you physically decrease protein adsorption

A

add steric hinderances

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8
Q

how do you decrease protein adsorption using steric hinderances

A

add hydrophilic, highly flexible chains on the surface that will physically block the proteins from coming into contact with the surface

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9
Q

for steric hinderance for protein adsorption, the chains must be __ because __

A

the chains must be hydrophilic so they can interact with the aqueous environment

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10
Q

what is PEG

A

PEG is a large, highly flexible hydrophilic polymer that decreases protein adsorption

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11
Q

PEG __ __ protein binding

A

decreases non specific protein binding

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12
Q

how does PEG decrease non specific protein binding

A

a large volume of the surface is taken up by these bulky chains that are in constant motion. This prevents protein adsorption

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13
Q

PEG is an example of ___ surface property that prevent protein adsorption

A

steric hinderance

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14
Q

what are the 4 ideal surface modification techniques

A

(TRDS)

  1. Thin (to minimize effects on bulk properties)
  2. Resistant to delamination (want it to stay on the surface)
  3. Simple and robust
  4. discourage surface rearrangement after treatment
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15
Q

what are examples of covalent surface coating

A

Plasma treatment and self assembling monolayers (SAM)

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16
Q

what is plasma treatment

A

assembly of species in a molecularly dissociated gaseous environment

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17
Q

what are the possible reactions of plasma treatment

A
  1. Etching - remove stuff from the surface
  2. Deposition - add stuff to the surface
  3. Functionaliztion - covalently add molecules
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18
Q

what is the name of the advantages of plasma treatment

A

Ratner and Hoffman

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19
Q

What the the advantages of plasma treatment

A

(g clefs)

  1. conformal - plasma will follow shape of implant
  2. free of voids
  3. easily prepared
  4. good adhesion to surface
  5. sterile
  6. low leachable substances
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20
Q

what are disadvantages of plasma treatment

A
  1. hard to predict composition of surface that will result
  2. expensive
  3. difficult to form uniform reaction
  4. easily contaminated
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21
Q

how are SAMs different than steric hinderance

A

SAMs have hydrophobic regions which will stabilize the structure (steric has hydrophilic chains)

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22
Q

why do SAMs attach to the surface

A

it is thermodynamically favored for them to attach and align

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23
Q

because they have a hydrophobic region, SAMs are __

A

amphiphilic

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24
Q

what are the 3 groups in SAMs

A
  1. Attachment group
  2. Long hydrocarbon chain
  3. Functional Group - can be used to alter hydrophobicity
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25
Q

SAMs result in __ reactions

A

exothermic

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26
Q

what is the purpose of the hydrophobic regions in SAMs

A

they stabilize the structure

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27
Q

what are advantages of SAMs

A
  1. ease of formation - can control specific areas where proteins will bind
  2. Chemically stable
  3. variety of functional groups are possible
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28
Q

disadvantages of SAMs

A

need for particular chemistry to drive attachment

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29
Q

what is an example of a non-covalent surface coating

A

surface modifying agents (SMA)

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30
Q

what are SMA

A

molecules that will spontaneously rise to the surface from the bulk, thus producing a coating

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31
Q

Unlike, plasma treatment and SAMs, SMA is a __

A

pre-fabrication design

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32
Q

what allows SMAs to form

A

there is a driving force to reduce the free energy at the surface

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33
Q

Which material is less likely to form SMAs and why

A

Ceramics because there is less atomic motion in the bulk due to electroneutrality

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34
Q

how do SMAs form in polymers

A

hydrophilic particles inside the polymer will want to move to the surface when the material is placed inside an aqueous environment

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35
Q

what determine the effectiveness of SMAs

A
  1. difference in free energy of the surface with and without the SMA
  2. Molecular motility of the SMA in the bulk to actually allow it to move to the surface
  3. Environment surrounding the implant must be hydrophilic to allow the hydrophilic SMA to rise to the surface
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36
Q

what do conversion coatings do (physiochemcial surface modification)

A

they form a passive layer on metals

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37
Q

Conversion coatings can spontaneously form due to __

A

the porbaix diagram

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38
Q

what is a major concern with biological surface modification techniques

A

maintaining the bioactivity of the biological molecules

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39
Q

PEG is an example of a __ for biological coatings

A

a spacer arm

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40
Q

in covalent biological coatings, a molecule can be bound to a __

A

spacer arm

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41
Q

how does a spacer arm work for biological coatings

A

the spacer arm provides space between the biomolecule and the surface. this allows for greater rotational freedom and improves the activity of the biomolecule

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42
Q

a spacer arm is an example of a __ biological coating

A

covalent

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43
Q

give an example of a noncovalent biological coating

A

a positively charged surface can attract heparin (which is negatively charged)

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44
Q

what information does contact angle analysis provide

A

information about the hydrophobicity of the surface

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45
Q

higher contact angel = __

A

more hydrophobic

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46
Q

what is hysteresis (for contact angle analysis)

A

surface tension of a material can be changed before and after water is added

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47
Q

contact angle analysis doesn’t provide any info about __

A

chemical composition of the surface

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48
Q

explain how contact angle analysis would appear with SMAs

A

SMA that would move surface in body is hydrophilic – surface becomes more hydrophilic with time – contact angle decreases with time, so advancing angle is larger

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49
Q

what technique would you use to view the 3D surface of a material

A

atomic force microscopy (ATF)

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50
Q

ATF doesn’t provide info about __

A

chemical composition

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51
Q

what technique would you use to find the chemical composition of a surface

A

ATR-FTIR

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52
Q

protein binding on hydrophilic surfaces are __

A

relatively reversible

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53
Q

protein binding on hydrophobic surfaces are __

A

irreversible (protein denature on surface)

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54
Q

at early time points, adsorption is ___ controled

A

diffusion

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55
Q

anything that lowers __ is favorable

A

∆G

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56
Q

what is the gibbs free energy equation for protein adsorption

A

∆Gadsorption = ∆Gprotein +∆Gsolvent + ∆Gsurface

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57
Q

adsorption will be thermodynamically favorable if __

A

the adsorption event lowers ∆G for one or more components without raising ∆G for another component

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58
Q

what factors have the largest impact on protein adsorption

A

(drs)

  1. dehydration of the surface and protein
  2. redistribution of charged groups
  3. structural rearrangment of proteins
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59
Q

explain how dehydration of the protein and surface affects protein adsorption

A

water molecules become more ordered at hydrophobic surfaces

if hydrophobic areas are able to come together through adsorption, they will reduce the hydrophobic surface area that is able to come into contact with water

this will allow water to become more disordered (increase in entropy/disorder is favored)

hydrophobic amino acids will come together to minimize its interactions with water

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60
Q

explain how redistribution of charge affects protein adsorptoin

A

adsorption is preferred when the material and the surface have opposite charges

in this case, the transfer of fewest ions is required

repulsion of material and surface can be overcome by adsorption of ions from the solution to help create opposite charges

but the adsorption of ions costs energy

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61
Q

explain how structural rearrangement of the protein affects protein adsorption

A

a less stable protein will adsorb more easily since conformational rearrangement is easier

allow for optimal conformation so that hydrophobic and charged regions of the protein can be placed to fulfill the two other properties (dehydration and redistribution of charged)

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62
Q

amino acids are considered __ because they can act as either ___

A

zwitterion - they can act as either acids or bases

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63
Q

what are the two amino acids that play a large role in determining 2º, 3º, and 4º structure

A

proline and cysteine

they are both non charged

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64
Q

what about proline and cysteine makes them important for determining 2º, 3º, and 4º structure

A

proline has a ring backbone that can limit protein folding in 3D

cysteine contains sulfide (SH) which can form disulfide bonds with other cysteine residues

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65
Q

__ is the linear order of amino acids

A

primary structure

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66
Q

__ is a disease in the

primary structure which dictates quaternary structure

A

sickle cell

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67
Q

__ and __ are examples of secondary structures

A

alpha helix and beta pleated sheets

68
Q

__ are localized interactions between amino acid residues

A

secondary structure

69
Q

what is the the 3D arrangement of an entire polypeptide chain

A

tertiary structure

amino acids on the same chain are interacting

70
Q

__ is an example of tertiary structure

71
Q

__ is the primary driving force for tertiary structure formation

A

hydrophobic interactions between non polar amino acids

minimize the amount of hydrophobic area that is interacting with the water

this is more energetically favorable (entropy)

72
Q

__ is the 3D arrangement of protein subunits

A

quaternary structure

interactions of amino acids between chains

73
Q

__ is an example of quaternary structure

A

hemoglobin

74
Q

there is a __ correlating to diffusion

A

high initial adsorption rate

75
Q

__ is reversible because the proteins are __

A

initial binding is reversible because proteins are not strongly bound to the surface

76
Q

after __, binding is irreversible

A

conformational changes

77
Q

why does a plateau occur in protein adsorption kinetics

A

a plateau occurs due to a monolayer coverage of the entire surface by proteins

78
Q

explain desorption and exchange

A

a protein with higher affinity for the surface will displace the protein already on the surface

79
Q

why is desorption unlikely after conformational changes occur in the protein

A

you would have to break all contacts between the surface and the protein

80
Q

what are the techniques for determining protein type and amount

A

HPLC and ELISA

81
Q

HPLC is used to determine __

A

the amount of protein in a mixture

82
Q

describe the different types of HPLC

A

(AANR)
adsorption chromatography - separation by adsorption based on hydrophobic and polar interactions. the first species to exit the column has least affinity for the column

normal phase chromatography - uses a polar stationary phase and a nonpolar mobile phase. more hydrophilic are eluted last since they have affinity for the polar column

reverse phase - hydrophobics elute last

affinity chromatography - integrate area under curve to find out how much of that species is present

83
Q

describe the process of an ELISA

A
  1. primary antibodies are added first
  2. the sample is then added
  3. the secondary antibody is added that will bind to the protein. the secondary antibody has an enzyme attached to it
  4. a substrate for the enzyme is added
    color change is proportional to the amount of protein present
84
Q

what does a modified ELISA tell?

A

used to determine the amount of protein already on the surface of a material

85
Q

how is modified ELISA different from an ELISA

A

in a modified ELISA, the protein is already attached to the surface. therefore you don’t need to used the primary antibodies normally used in an ELISA

86
Q

a negative result in an ELISA can mean__

A

there is no protein or the protein is denatured

87
Q

__ have attained tissue -specific functions

A

differentiated cells

88
Q

__ can differentiate into a variety of cell types

A

progenitor cells

89
Q

what are proliferation assays?

A

that are basically viability assays over time

90
Q

when cell spreading, __ interacts with __ to anchor the cell firmly in place

A

the integrin interacts with the ligands

91
Q

describe the process of cell migration

A
  1. pseudopodia extend
  2. membrane attaches to the substrate via integrin
  3. after pseudopodia are adhered, there is a contractile force and the release of rear receptors
  4. integrin receptors are recycled from back to front
92
Q

__ is the speed of cell movement

A

translocation speed

93
Q

__ is the time the cell moves without changing direction

A

persistance time

94
Q

what are the cytotoxicity assays

A

Direct contact assays
Agar diffusion assay
elution assay

95
Q

in a __, the material is placed over top of the cells and cells death in the surrounding area is observed

A

direct contact assay

96
Q

in a direct contact assay, you need __

A

both a positive (non-cytotoxic) and negative (cytotoxic) control

97
Q

what is an agar diffusion assay

A

cells are covered with agar before the material is placed on top

soluble products from material diffuse into the agar

98
Q

describe the results seen from an agar diffusion assay

A

if cell death occurs farther away from the sample, the more cytotoxic the sample

99
Q

__ are performed to determine the cytotoxicity of leachable molecules found in biomaterials

A

elution assays

100
Q

describe how a adhesion/spreading assay works

A
  1. allow cells to adhere to surface then wash away non-adhered cells
  2. number of cells -on suface = total number of cells - cells washed away
101
Q

what are the disadvantages of cell adhesion/spreading assays

A

concerns over reproducibility of the force exerted in the washing step

102
Q

what are the migration assays

A

capillary tube test

boyden chamber assay

103
Q

describe a capillary tube test

A
  1. confine cell population in a small tube.
  2. the opening of a tube is placed against a surface, allowing the cells to migrate out in a fan like shape
  3. the area of the fan shape is measured
104
Q

disadvantages of capillary tube test

A

it is difficult to distinguish the effects of proliferation and migration

105
Q

what does a boyden chamber assay tell

A

looks at how cells might migrate in response to materials that are leached
(cells might move closer to the leached material)

106
Q

this assay is used to determine the effects of soluble factors on migration rather than examining the impact of the substrate

A

boyden chamber assay

107
Q

how does a boyden chamber assay work

A
  1. filter separates the cells from the soluble agents
  2. cells are seeded on the top layer and the substance is on the bottom layer
  3. cells will move to the bottom chamber if they are attracted to the leached materials
108
Q

how do you track individual cells

A
  1. put cells on material
  2. take pictures to determine where they move over time

speed of migration
persistence time

109
Q

speed of migration and persistence time are associated with __

A

tracking individual cells

110
Q

longer persistence time =

A

chemotactic response

111
Q

what type of immunity is always present

A

innate immunity

112
Q

__ provides the first line of defense

A

innate immunity

113
Q

if the invading organism is not cleared by the innate immune response, then the body engages the _

A

acquired immune response

114
Q

what are the components of the innate immune response

A
  1. barriers (skin)
  2. physiologic barriers (pH in stomach)
  3. Phagocytic cells
  4. inflammation
115
Q

what are the two types of inflammation

A

acute

chronic

116
Q

__ occurs immediately after an injury

A

acute inflammation

117
Q

of the two types of inflammation, which one is undesired

A

chronic inflammation

118
Q

__ inflammation persists over weeks to months

A

chronic inflammation

119
Q

redness, swelling, heating, pain are signs of __

A

acute inflammation

120
Q

Acquired immunity involves the activation of __

A

white blood cells called lymphocytes

121
Q

acquired immunity responses to a particular _

122
Q

when does infection develop2

A

if the invader persists after exposure to both the innate and acquired immune response

123
Q

red blood cells are known as

A

erythrocytes

124
Q

lymphocytes are part of the __

A

acquired immune response

125
Q

granulocytes are __

A

phagocytic cells in innate immunity

126
Q

neutrophils are a type of ___

A

granulocytes

127
Q

what is the first step in the timeline of an injury (acute inflammation)

A

tissue macrophages release cytokines to call in neutrophils (chemotaxis)

128
Q

after tissue macrophages call neutrophils, what happens in the timeline for acute inflammation

A

neutrophils arrive and move out of the blood and into the tissue via extravasation

129
Q

describe the process for extravasation

A
  1. rolling - selectins are upregulated in inflamed endothelium. the selectins interact with the neutrophils which helps bind them to the endothelium
  2. activation - neutrophils are activated by cytokines that interact with integrins on the neutrophils. once activated, the integrins have higher affinity for CAMs on the endothelium
  3. Arrest and adhesion - strong interactions between integrin on neutrophils and CAM on endothelium cause the neutrophil to adhere to the endothelium
  4. Migration - diapedesis, the neutrophils squeezes through the endothelium cells and into the tissue
130
Q

after extravasation, what do the neutrophils do in acute inflammation

A

once in the itssue, the neutrophils will phagocytose

they can undergo respiratory burst - create reactive oxygen species to kill invaders

they secrete cytokines that recruit monocytes

they can secrete cytokine that release factors that are chemotactic for lymphocytes
**provides a point of communication or crossover between the innate and acquired immune system **

131
Q

after the neutrophils secrete cytokines to recruit monocytes, what happens

A

monocytes arrive and become macrophages

132
Q

how do macrophages work

A

phagocytosis

respiratory burst

secrete cytokines to increase acquired immune response

act as antigen presenting cells*

133
Q

describe a potential problem during phagocytosis by macrophages

A

small particles - macrophages will eat non degradable particles. but the particle will be re-released when the macrophage dies (silicosis)

large particles - large particles undergo frustrated phagocytosis. the macrophage will release lysosomal enzyme outside of the cell to digest the invader. too much of this can lead to tissue damage

134
Q

how to terminate acute inflammation

A

short half life of chemical mediators

cytokines are produced that can act as inhibitors to pro-inflammatory cytokines

135
Q

what are the types of cells that can be used for assaying acute immune response

A

leukocytes

endothelial cells

136
Q

leukocyte cell assays are used for assaying what type of cells

A

neutrophils and macrophages

137
Q

what can you determine using leukocytes cells in an assay

A

adhesion/spreading - more spreading means more inflammatory

cell migration - more migration means more inflammatory (boyden chamber assay)

pro inflammatory cytokine release - use ELISA to detect pro inflammatory cytokines

look at cell surface markers - use FACS (similar to ELISA but works on cell surface) increase in markers/ receptors means more inflammatory

138
Q

what can you determine using endothelial cell in an assay to

A

increase in cell surface receptors on endothelial cells (selectin, CAMs) means more inflammation

This promotes the migration of neutrophils

139
Q

1-3 days after the injury, macrophages call in __ to __

A

call in fibroblasts and vascular endothelial cells

140
Q

proliferation of fibroblasts and vascular endothelial cells leads to the formation of __

A

granulation tissue

141
Q

granulation tissue may form part of the __

A

foreign body reaction

142
Q

what are foreign body giant cells (FBGCs)

A

they are fused macrophages which form in an attempt to phagocytose the biomaterial, which is much larger than a single cell

143
Q

why does foreign body giant cells form

A

to attempt to phagocytose biomaterials which are much larger than a single macrophage

144
Q

what type for foreign body reaction will the following materials get:

smooth implants

rough implants

high surface area to volume (porous)

Low surface area to volume

A

smooth - get macrophages

rough - get macrophages and FBGCs

high SA - marcophages and FBGCs

low SA - fibrous (granulation tissue)
granulation tissue has more collagen and has higher mechanical properties

145
Q

how long do FBGCs remain

A

they may remain for the lifetime of the implant but not sure if they are actively secreting enzymes the entire time

146
Q

fibrous encapsulation only pertains to what type of biomaterials

A

non-degradable

147
Q

what is fibrous encapsulation

A

it is the maturation of granulation tissue, which is marked by the presence of large blood vessels and collagen

happens around an implant

148
Q

the thickness of fibrous encapsulation depends on __

A

the degree of injury

location of implant - more motion can lead to thicker encapsulation

shape - edges leads to thicker capsule

degradation - short degradation will lead to no capsule

149
Q

in chronic inflammation __ has to be there for a long time

A

lymphocytes

150
Q

you see __ in chronic inflammation

A

granulomas - consists of layers of FBGCs surrounding non-phagocoytosable particles

151
Q

what are the 4 resolutions after implantation

A
  1. encapsulation
  2. integration - no fibrous layer, cells from tissue form right next to the implant
  3. resorption - if the implant degrades, no capsule will form
  4. extrusion - the implant will be forced out
152
Q

repair vs. regeneragtion

A

repair - the defect is replaced with scar tissue (occurs when cut extends into dermal layer)

regeneration - identical tissue is formed (only occur when injury is in epithelial layer )

153
Q

how to do you tell if something is repair or regeneration in vivo?

A

you can’t. you need to take a sample of the tissue and test it in vitro

154
Q

what are the consideration for in vivo studies for implants

A
  1. choice of animal
  2. implant site
  3. length of study
  4. biomaterial considerations (shape, size)
  5. controls
155
Q

what are the types of controls used in studies

A

contralateral control (opposite uninjured limb) shows level of regeneration

unfilled defect - asses level of improved healing

156
Q

what is responsible for the allergic response

A

acquired immune response

157
Q

what are the 4 characteristics of acquired immunity

A
  1. specificity - antigen will bind to specific antibody in the humoral response
  2. diversity - response to a wide array of thing
  3. self/non self recognition - problem with autoimmune disorders
  4. memory - this is why vaccines work
158
Q

what are the 2 types of acquired immunity

A

humoral immunity - action of antibodies

cellular immunity - action of t cells

159
Q

describe humoral immunity

A

it is the action of antibodies. mediated by B cells

main way to fight off foreign invaders

160
Q

describe cellular immunity

A

action of t cells

detect altered self cells

161
Q

the specificity of acquired immunity comes from __

A

lymphocytes (b and t cells) that are involved in an immune reaction are active in response to antigens

162
Q

what is an antigen

A

it is a substance that binds to an antibody or t cell receptor to initiate the acquired immune response

antigen has a high molecular weight

163
Q

what is a hapten

A

it is a low MW substance that combines with a high MW molecule to induce an immune response

upon subsequent exposure, you don’t need the larger molecule to induce an immune response

the small molecule can induce an immune response by itself even though it normally wouldn’t (metal allergies)

164
Q

what is an adjuvant

A

it non specifically enhances immune response to antigens

it increases uptake of antigens by phagocytic cells

165
Q

what does the compliment system do

A

it causes the elimination of foreign elements

it is the bridge between innate immunity and acquired immunity

166
Q

what are the 2 parts of the compliment system

A

the classical and alternative pathway.

they converge to cause formation of the membrane attack complex to lyse foreign cells

167
Q

how does the classical pathway begin

A

with the binding of an antibody to an antigen on a bacterial cell

the attachment of the antibody induces a conformational change, which causes a cascade of chemical reactions starting with the cleavage of C4

BMED 4751 (3 decks)

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