Background
Methods
Animals
Cohort | Age (months) | Group | n | Sample Type | Analyses Performed |
---|---|---|---|---|---|
1* | 12 | Adult | 5 | ||
28 | Aged Intact | 8 | synaptosomes (whole hippocampus) | Morris water maze, transcriptomic analysis, qPCR | |
28 | Aged Impaired | 7 | |||
2 | 12 | Adult | 7 | ||
26 | Aged Intact | 7 | Hippocampal subregions (CA1, CA3, DG) | Morris water maze, qPCR, immunoblotting | |
26 | Aged Impaired | 10 | |||
3* | 13 | Adult | 3 | ||
26 | Aged Intact | 3 | Perfusion-fixed sagittal brain sections | Morris water maze, immunohistochemistry | |
26 | Aged Impaired | 3 |
Morris water maze testing
Synaptosome isolation
Hippocampal subregion dissection
Perfusion fixation and embedding
RNA isolation
Microarray analysis
Bioinformatic analysis and visualization
RT-qPCR
Gene Symbol | Gene ID | Gene Name | TaqMan AOD # |
---|---|---|---|
C1s | 192262 | complement component 1, subcomponent s | Rn00594278_m1 |
C3 | 24232 | complement component 3 | Rn00566466_m1 |
C4a | 24233 | complement component 4a | Rn00709527_m1 |
Ccr5 | 117029 | chemokine (C-C motif) receptor 5 | Rn00588629_m1 |
Cd74 | 25599 | CD74 molecule, major histocompatibility complex, class II invariant chain | Rn00565062_m1 |
Cp | 24268 | ceruloplasmin | Rn00561049_m1 |
Ctse | 25424 | cathepsin E | Rn00564036_m1 |
Erbb3 | 29496 | v-erb-b2 erythroblastic leukemia viral oncogene homolog 3 | Rn00568107_m1 |
Fcgr2a | 116591 | Fc fragment of IgG, low affinity IIa, receptor (CD32) | Rn00821543_g1 |
Fcgr2b | 289211 | Fc fragment of IgG, low affinity IIb, receptor (CD32) | Rn00598391_m1 |
Hla-dmb | 3109 | major histocompatibility complex, class II, DM beta | Rn01429041_m1 |
Hla-dra | 294269 | major histocompatibility complex, class II, DR alpha | Rn01427980_m1 |
Hla-drb1 | 294270 | major histocompatibility complex, class II, DR beta 1 | Rn01429350_m1 |
Hsbp1 | 286899 | heat shock factor binding protein 1 | Rn00583001_g1 |
Icam1 | 25464 | intercellular adhesion molecule 1 | Rn00564227_m1 |
Lgals3 | 83781 | lectin, galactoside-binding, soluble, 3 | Rn00582910_m1 |
S100a6 | 85247 | S100 calcium binding protein A6 | Rn00821474_g1 |
Serping1 | 295703 | serpin peptidase inhibitor, clade G (C1 inhibitor), member 1 | Rn01485600_m1 |
Tlr2 | 310553 | toll-like receptor 2 | Rn02133647_s1 |
Tlr4 | 29260 | toll-like receptor 4 | Rn00569848_m1 |
Tlr7 | 317468 | toll-like receptor 7 | Rn01771083_s1 |
Protein Extraction
Immunoblotting
Target | Supplier | Catalog # | Host | Use | Method* |
---|---|---|---|---|---|
Glial fibrillary acidic protein (GFAP) | Abcam | 7620 | rabbit | primary | IB/IHC |
Ionized calcium binding adaptor molecule 1 (Iba1) | Wako Pure Chemical Industries | 1620001 | rabbit | primary | IB |
Ionized calcium binding adaptor molecule 1 (Iba1) | Wako Pure Chemical Industries | 1919741 | rabbit | primary | IHC |
MHC2 invariant chain (CD74; Ox6) | Abcam | 23990 | mouse | primary | IHC |
Rabbit IgG (HRP-conjugated) | GE Healthcare | NA934V | donkey | secondary | IB |
Rabbit [F(ab')2] | Jackson ImmunoResearch | 711496152 | donkey | secondary | IHC |
Mouse [F(ab')2] | Jackson ImmunoResearch | 715486150 | goat | secondary | IHC |
Immunohistochemistry
Quantitation of astrocyte and microglia populations
Results
Behavioral stratification of adult and aged rats by Morris water maze performance
Age-related upregulation of the MHC II antigen presentation pathway
Differential MHC II and related gene expression with aging occurs throughout the hippocampus
Gene | All Aged vs. Adult (ratio, p-value t-test)
| Pearson Correlation to Morris water maze | Aged Intact vs. Adult (ratio, p-value SNK)
| Aged Impaired vs. Adult(ratio, p-value SNK)
| Aged Impaired vs. Aged Intact(ratio, p-value SNK)
| |||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Syn | CA1 | CA3 | DG | Syn | CA1 | CA3 | DG | Syn | CA1 | CA3 | DG | Syn | CA1 | CA3 | DG | Syn | CA1 | CA3 | DG | |
MHC II Components
| ||||||||||||||||||||
Cd74
| 6.58###
| 5.61###
| 3.61###
| 2.43###
| - | - | - | - | 6.79*** | 5.66*** | 3.88*** | 2.43*** | 6.35*** | 5.58*** | 3.40*** | 2.49*** | - | - | - |
-
|
Ctse
| 1.69##
| 1.45##
| 1.75###
| 1.48###
| - | - | - | - | 1.81* |
-
| 1.73*** | 1.48* |
-
|
-
| 1.77*** | 1.54** | - | - | - |
-
|
Hla-dmb
| 1.64###
| 1.37###
| 1.37###
| 1.26##
| - | - | - | - | 1.57** | 1.33** | 1.39*** |
-
| 1.72*** | 1.40*** | 1.35*** | 1.33** | - | - | - | 1.16* |
Hla-dra
| 6.30###
| 4.04###
| 3.02###
| - | - | - | - | - | 6.76*** | 4.18*** | 3.25*** |
-
| 5.75** | 3.95*** | 2.85*** |
-
| - | - | - |
-
|
Hla-drb1
| 10.50###
| 7.13###
| 4.39###
| 2.91#
| - | - | - | - | 10.70*** | 7.12*** | 4.77*** |
-
| 10.26*** | 7.13*** | 4.12*** | 3.42** | - | - | - |
-
|
Antigen Recognition
| ||||||||||||||||||||
C1s
| 1.84#
| 1.21#
| 1.56###
| - | - | - | - | - | - |
-
| 1.59** | - | - |
-
| 1.53*** | - | - | - | - |
-
|
C3
| 3.47###
| 2.00###
| 1.96###
| 1.69###
| - | - | - | - | 3.40** | 2.07*** | 2.22*** | 1.52*** | 3.55** | 1.94*** | 1.78*** | 1.81*** | - | - | - | 1.19* |
C4a
| 4.18###
| 1.77###
| 1.51###
| - | - | - | - | - | 3.83** | 1.67*** | 1.58*** |
-
| 4.59** | 1.85*** | 1.47*** |
-
| - | - | - |
-
|
Serping1
| 2.71###
| 2.03###
| 2.27###
| 1.54###
| - | - | - | - | 2.79*** | 2.18*** | 2.52*** | 1.49** | 2.63*** | 1.92*** | 2.09*** | 1.58*** | - | - | - |
-
|
Tlr2
| 1.64##
|
-
| 1.54## | - | - | - | - | - | 1.55* |
-
| 1.66* |
-
| 1.73* |
-
| 1.46* |
-
| - | - | - |
-
|
Tlr4
| 1.51##
| 1.35#
| 1.96###
| - | - | - | - | - | 1.41* |
-
| 2.00*** |
-
| 1.60* |
-
| 1.93*** |
-
| - | - | - |
-
|
Tlr7
| 1.69##
| 1.34##
| 1.73###
|
-
| - | - | - | - | 1.65* | 1.34* | 1.74*** |
-
| 1.73* | 1.35* | 1.72*** |
-
| - | - | - |
-
|
Immune Cell Activating
| ||||||||||||||||||||
Ccr5
| 1.96#
|
-
| - |
-
| - | - | - | - | - |
-
| - |
-
|
-
|
-
|
-
|
-
| - | - | - |
-
|
Erbb3
| 1.28###
| 1.34###
| 1.20#
| 1.40#
| - | - | - | - | 1.26** | 1.43*** | - |
-
| 1.30** | 1.28** |
-
| 1.47** | - | - | - |
-
|
Fcgr2a
| 1.45#
| 1.19#
| 1.30###
| - | - | - | - | - |
-
| 1.30** |
-
|
-
|
-
| 1.30** |
-
| - | - | - |
-
| |
Fcgr2b
| 1.96##
|
-
| 1.61###
|
-
| - | - | - | - | 2.08* |
-
| 1.63*** |
-
| 1.82* |
-
| 1.60*** |
-
| - | - | - |
-
|
Inflammation Response
| ||||||||||||||||||||
Cp
| 1.67#
|
-
| 1.56###
|
-
| - | - | - | - | 1.92* |
-
| 1.64*** |
-
|
-
|
-
| 1.50*** |
-
| - | - | - |
-
|
Hsbp1
| 2.87###
| 1.68###
| 2.09###
|
-
| - | - | - | - | 3.13** | 1.65** | 2.25*** |
-
| 2.58*** | 1.70** | 1.97*** |
-
| - | - | - |
-
|
Icam1
| 1.46#
| 1.55###
| 1.43###
|
-
| - | - | - | - |
-
| 1.68*** | 1.64*** |
-
|
-
| 1.46** | 1.31** |
-
| - | - | - |
-
|
Lgals3
| 3.42##
| 2.64###
| 1.98###
| 1.84##
| - | - | - | - | 3.68*** | 2.62*** | 2.18*** | 1.63* | 3.03*** | 2.66*** | 1.84*** | 2.00** | - | - | - |
-
|
S100a6
| 1.52#
| 2.03###
| 2.21###
|
-
| - | - | - | - |
-
| 2.00*** | 2.48*** |
-
| - | 2.06*** | 2.02*** |
-
| - | - | - |
-
|
CA1 | CA3 | DG | synaptosomes (whole-hippocampus) | |||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Aged
|
Aged
|
Aged
|
Aged
|
Aged
|
Aged
|
Aged
|
Aged
| |||||||||
Gene
|
Adult
|
Aged
|
Intact
|
Impaired
|
Adult
|
Aged
|
Intact
|
Impaired
|
Adult
|
Aged
|
Intact
|
Impaired
|
Adult
|
Aged
|
Intact
|
Impaired
|
C1s
|
100 ± 2.8
|
121 ± 6.1
| 125 ± 11.7 | 118 ± 6.6 |
100 ± 7.8
|
156 ± 7.6
| 159 ± 13.4 | 153 ± 9.3 |
100 ± 6.1
|
122 ± 4.8
| 117 ± 5.7 | 125 ± 6.9 |
100 ± 21.7
|
184 ± 21.8
| 182 ± 28.6 | 186 ± 35.3 |
C3
|
100 ± 6.6
|
200 ± 10.9
| 207 ± 16.4 | 194 ± 15.0 |
100 ± 7.9
|
196 ± 10.8
| 222 ± 17.8 | 178 ± 10.7 |
100 ± 8.1
|
169 ± 7.2
| 152 ± 10.2 | 181 ± 8.3 |
100 ± 23.2
|
347 ± 35.7
| 340 ± 27.6 | 355 ± 74.4 |
Serping1
|
100 ± 7.4
|
203 ± 13.4
| 218 ± 14.6 | 192 ± 20.2 |
100 ± 7.3
|
227 ± 11.5
| 252 ± 17.4 | 209 ± 13.1 |
100 ± 4.8
|
154 ± 6.3
| 149 ± 9.4 | 158 ± 8.7 |
100 ± 6.9
|
271 ± 12.4
| 279 ± 19.9 | 263 ± 15.5 |
Cd74
|
100 ± 7.7
|
561 ± 33.3
| 566 ± 44.5 | 558 ± 47.9 |
100 ± 10.9
|
361 ± 20.5
| 388 ± 37.6 | 340 ± 21.2 |
100 ± 7.7
|
243 ± 17.2
| 233 ± 36.7 | 249 ± 18.2 |
100 ± 28.8
|
658 ± 48.9
| 679 ± 46.7 | 635 ± 95.8 |
C4a
|
100 ± 7.1
|
177 ± 7.0
| 167 ± 11.1 | 185 ± 8.6 |
100 ± 4.8
|
151 ± 4.6
| 158 ± 6.2 | 147 ± 6.2 |
100 ± 9.7
|
135 ± 10.3
| 117 ± 15.2 | 147 ± 13.0 |
100 ± 17.7
|
418 ± 44.6
| 383 ± 31.6 | 459 ± 88.3 |
Ccr5
|
100 ± 5.2
|
108 ± 5.1
| 106 ± 5.2 | 110 ± 7.8 |
100 ± 3.1
|
121 ± 6.3
| 117 ± 7.3 | 125 ± 10.3 |
100 ± 6.9
|
99 ± 7.0
| 88 ± 7.1 | 106 ± 10.4 |
100 ± 17.9
|
196 ± 23.0
| 172 ± 18.8 | 220 ± 41.8 |
Cp
|
100 ± 5.4
|
93 ± 8.2
| 93 ± 6.9 | 93 ± 13.5 |
100 ± 3.9
|
156 ± 6.3
| 164 ± 9.2 | 150 ± 8.4 |
100 ± 9.2
|
110 ± 8.0
| 118 ± 13.7 | 103 ± 9.8 |
100 ± 10.8
|
167 ± 17.1
| 192 ± 28.6 | 146 ± 18.4 |
Ctse
|
100 ± 8.0
|
145 ± 10.2
| 141 ± 13.7 | 148 ± 14.4 |
100 ± 9.8
|
175 ± 7.4
| 173 ± 13.9 | 177 ± 8.7 |
100 ± 6.8
|
148 ± 6.8
| 140 ± 11.3 | 154 ± 8.4 |
100 ± 21.0
|
169 ± 12.0
| 181 ± 19.8 | 156 ± 11.4 |
Erbb3
|
100 ± 2.7
|
134 ± 5.1
| 143 ± 7.5 | 128 ± 6.5 |
100 ± 8.1
|
120 ± 5.1
| 128 ± 9.1 | 114 ± 5.6 |
100 ± 7.5
|
140 ± 7.8
| 129 ± 10.9 | 147 ± 10.3 |
100 ± 5.4
|
128 ± 4.0
| 126 ± 6.5 | 130 ± 4.6 |
Fcgr2b
|
100 ± 3.2
|
108 ± 7.9
| 103 ± 6.4 | 112 ± 13.0 |
100 ± 9.0
|
161 ± 5.2
| 163 ± 12.0 | 160 ± 3.6 |
100 ± 4.2
|
124 ± 11.0
| 125 ± 19.8 | 123 ± 13.2 |
100 ± 20.8
|
196 ± 15.4
| 208 ± 22.6 | 182 ± 21.2 |
Fcgr2a
|
100 ± 4.4
|
119 ± 4.8
| 116 ± 3.8 | 120 ± 7.7 |
100 ± 5.2
|
130 ± 4.2
| 130 ± 6.4 | 130 ± 5.9 |
100 ± 5.1
|
115 ± 3.8
| 109 ± 5.0 | 120 ± 5.0 |
100 ± 15.2
|
145 ± 11.4
| 143 ± 10.7 | 147 ± 22.8 |
Hla-dmb
|
100 ± 4.6
|
137 ± 4.6
| 133 ± 5.2 | 140 ± 7.3 |
100 ± 3.9
|
137 ± 3.7
| 139 ± 5.9 | 135 ± 4.9 |
100 ± 7.4
|
126 ± 4.4
| 115 ± 6.6 | 133 ± 4.7 |
100 ± 3.3
|
164 ± 8.3
| 157 ± 13.7 | 172 ± 8.4 |
Hla-dra
|
100 ± 7.9
|
404 ± 19.0
| 418 ± 28.1 | 395 ± 26.5 |
100 ± 14.7
|
302 ± 23.4
| 325 ± 47.0 | 285 ± 23.4 |
100 ± 9.7
|
181 ± 22.4
| 190 ± 43.4 | 175 ± 26.4 |
100 ± 24.0
|
630 ± 51.8
| 676 ± 67.9 | 575 ± 79.8 |
Hsbp1
|
100 ± 12.8
|
168 ± 9.6
| 165 ± 14.0 | 170 ± 13.6 |
100 ± 8.7
|
209 ± 11.0
| 225 ± 18.7 | 197 ± 13.0 |
100 ± 9.5
|
109 ± 11.5
| 80 ± 14.2 | 130 ± 13.8 |
100 ± 17.7
|
287 ± 25.1
| 313 ± 30.1 | 258 ± 40.9 |
Icam1
|
100 ± 5.5
|
155 ± 7.6
| 168 ± 11.9 | 146 ± 9.4 |
100 ± 5.6
|
143 ± 4.2
| 164 ± 13.3 | 131 ± 5.7 |
100 ± 12.2
|
102 ± 6.5
| 100 ± 13.5 | 102 ± 6.5 |
100 ± 11.1
|
146 ± 11.8
| 145 ± 16.7 | 147 ± 18.3 |
Lgals3
|
100 ± 4.9
|
264 ± 16.6
| 262 ± 21.4 | 266 ± 24.9 |
100 ± 7.3
|
198 ± 12.6
| 218 ± 16.3 | 184 ± 17.4 |
100 ± 10.5
|
184 ± 12.9
| 163 ± 10.0 | 200 ± 20.2 |
100 ± 21.4
|
342 ± 34.1
| 368 ± 54.6 | 303 ± 39.2 |
Hla-drb1
|
100 ± 11.0
|
713 ± 57.0
| 712 ± 96.9 | 713 ± 73.8 |
100 ± 9.7
|
439 ± 34.8
| 477 ± 73.9 | 412 ± 30.1 |
100 ± 14.1
|
291 ± 40.2
| 225 ± 44.4 | 342 ± 59.0 |
100 ± 7.6
|
1050 ± 67.3
| 1070 ± 97.4 | 1026 ± 100.2 |
S100a6
|
100 ± 2.8
|
203 ± 11.5
| 200 ± 16.2 | 206 ± 16.9 |
100 ± 7.5
|
221 ± 13.4
| 248 ± 20.9 | 202 ± 15.5 |
100 ± 16.3
|
110 ± 8.3
| 88 ± 15.5 | 123 ± 7.4 |
100 ± 9.0
|
152 ± 13.1
| 161 ± 24.3 | 145 ± 15.5 |
Tlr2
|
100 ± 6.3
|
106 ± 5.7
| 109 ± 9.8 | 104 ± 7.2 |
100 ± 11.2
|
154 ± 9.6
| 166 ± 18.9 | 146 ± 9.7 |
100 ± 16.6
|
172 ± 18.8
| 166 ± 36.7 | 176 ± 22.7 |
100 ± 13.0
|
164 ± 11.3
| 155 ± 19.2 | 173 ± 12.9 |
Tlr4
|
100 ± 2.7
|
135 ± 8.1
| 118 ± 12.8 | 143 ± 9.7 |
100 ± 8.4
|
196 ± 9.5
| 200 ± 19.7 | 193 ± 9.6 |
100 ± 8.7
|
119 ± 4.0
| 118 ± 5.9 | 120 ± 5.7 |
100 ± 8.3
|
151 ± 10.1
| 141 ± 4.9 | 160 ± 19.7 |
Tlr7
|
100 ± 3.5
|
134 ± 6.5
| 134 ± 17.2 | 135 ± 3.9 |
100 ± 8.1
|
173 ± 5.0
| 174 ± 9.6 | 172 ± 5.7 |
100 ± 11.6
|
94 ± 7.6
| 95 ± 11.6 | 93 ± 10.7 |
100 ± 13.3
|
169 ± 11.3
| 165 ± 16.6 | 173 ± 16.5 |