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Pathobiology of the senescence-accelerated mouse (SAM)
ExperimentalGerontology,Vol. 32, Nos. I/2,pp. 117-127, 1997 Copyright© 1997ElsevierScienceInc. Printedin the USA.All rightsreserved 0531-5565/97$17.00+ .00 ELSEVIER
PII S0531-5565(96)00068-X
PATHOBIOLOGY OF THE SENESCENCE-ACCELERATED MOUSE (SAM)
TOSHIO TAKEDA,l TAKATOSHIMATSUSHITA,2 MAFUMIKUROZUMI,3 KIMINOBUTAKEMURA,2 KEIICHI HIGUCHI2 and MASANORIHOSOKAWA2 IThe Council for SAM Research, Departmentof Senescence Biology and 2Departmentsof Senescence Biology and 3ClinicalLaboratory, Chest Disease Research Institute,Kyoto University,Sakyo-ku,Kyoto606, Japan
Abstract--Routine postmortem examinations and the pathobiological features revealed by systematically designed studies have shown several pathologic phenotypes that are often characteristic enough to differentiate among the various SAM strains: senile amyloidosis in SAMP1, -P2, -P7, -P9, -P10, and -P11; secondary amyloidosis in SAMP2 and -P6; contracted kidney in SAMP1, -P2, -P10, and - P l l ; immunoblastic lymphoma in SAMR1 and -R4; histiocytic sarcoma in SAMR1 and -R4; ovarian cysts in SAMR1; impaired immune response in SAMP1, -P2, and -P8; hyperinflation of the lungs in SAMP1; hearing impairment in SAMPI; degenerative temporomandibular joint disease in SAMP3; senile osteoporosis in SAMP6; deficits in learning and memory, in SAMP8 and -P10; emotional disorders in SAMP8 and -P10; cataracts in SAMP9; and brain atroFhy in SAMP10. These are all age-associated pathologies, the .incidence and severity of which increase with advancing age. The SAM model in which these pathobiological features have been carefully monitored will be a valuable tool in the clarification of the pathogenic mechanisms of age-associated pathologies and in the research for effective methods to modnLate or ameliorate these pathologies. Copyright © 1997 Elsevier Science Inc.
Key Words: Senescence-Accelerated Mouse, SAM, aging, senescence, pathobioiogy, pathologic phenotypes, age-associated pathologies, autopsy findings, postmortem examination INTRODUCTION SENESCENCE-ACCELERATED Mouse (SAM) has been under development at Kyoto University since 1970 by selective sister-brother matings of mice of the AKR/J strain donated by the Jackson Laboratory (Bar Harbor, ME) in 1968. At present, there are 12 lines: senescence-prone inbred strains (SAMP) include S A M P I , SAMP2, SAMP3; SAMP6, SAMP7, SAMP8, SAMP9, SAMP10, and S A M P l l ; and senescence-resistant strains (SAMR) SAMR1, SAMR4, and SAMR5 (Takeda et al., 1981, 1991, 1994).
Correspondence to: Toshio Takeda, The Council for SAM Research, Department of Senescence Biology, Chest Disease Research Institute, Kyoto University,Sakyo-ku, Kyoto 606, Japan 117
I 18
T. TAKEDA
e¢ al
In the first report on SAM (Takeda et al., 1981), amyloid deposition, abscess, thymic and nonthymic lymphoma, and pneumonia were cardinal autopsy findings in both the R and the P series, progenitors of SAMR and SAMP. The pathologic findings of recent routine postmortem examinations of SAMP and SAMR strains are described in this report, and the causes of death are analyzed. In addition, a series of extensive studies on the nervous, skeletal, sensory, respiratory, and immune systems have been conducted on living as well as dead SAM mice in a more dynamic approach to age-related pathobiological changes. Our findings include many pathobiological features, some of which are specific for each strain of the SAM model. AUTOPSY FINDINGS OF ROUTINE POSTMORTEM EXAMINATIONS Five hundred and sixty-five SAMP mice, 291 males and 274 females (62, 76, 66, 49, 88, 46, 35, 102, and 41 SAMP1, -P2, -P3, -P6, -P7, -P8, -P9, -P10, and -PI 1, respectively) and 204 SAMR mice, 103 males and 101 females (77, 52, and 75 SAMR1, -R4, and -R5, respectively), including both fresh carcasses and mice killed while moribund, were examined grossly and histologically. All these mice were reared under conventional conditions at the Chest Disease Research Institute, Kyoto University. The organs examined were the trachea, tongue, pharynx, larynx, salivary glands, thymus, thyroid, liver, kidney, spleen, lungs, heart, aorta, adrenal glands, abdominal skin, esophagus, alimentary tract, pancreas, and reproductive organs. As a rule, other organs and tissues, such as brain, spinal cord, nasal cavity, middle and inner ear, skeletal tissues, lymph nodes, etc., were not examined routinely. All organs and tissues were fixed in 10% buffered formalin and embedded in paraffin. The stains used were hematoxylin-eosin (4 Ia,m sections) and alkaline Congo red (6 ~m sections). When necessary, special stains were used: fat, PTAH (phosphotungstic acid hematoxylin), Azan-Mallory, and periodic acid-Schiff (PAS) or immunohistochemical stains with anti-apoA-II and anti-AA antisera for the identification of amyloid protein (Higuchi et al., 1983, 1991a,b). The gross and microscopic pathomorphological findings are summarized in Table 1, and the incidence of each pathologic finding is listed. The definite presence of any abnormality was considered positive, regardless of its severity. For example, the incidence of amyloidosis indicates the presence of amyloid definitely observed in any organ or tissue regardless of the severity of amyloid deposition. The autopsy findings were classified according to the type of change: inflammation, including bronchopneumonia, interstitial pneumonia, abscess, colitis, and other inflammatory changes; amyloidosis; contracted kidney; degeneration of renal tubules; invagination; angionecrosis; angiitis; lymphoid cell neoplasms, including thymic lymphoblastic lymphoma, nonthymic lymphoblastic lymphoma, immunoblastic lymphoma, follicular center cell lymphoma, and unclassifiable lymphomas; histiocytic sarcoma; granulocytic leukemia; and other neoplasms, including malignant neoplasms, ovarian cysts, and other benign neoplasms, such as hemangioma and pulmonary adenoma. Other miscellaneous pathologies were observed sporadically, such as pulmonary edema, hyperplasia of the extrahepatic bile duct, etc., but they are not listed. Interstitial pneumonia was often observed in mice with an abscess in any organ or tissue, but the lesions did not advance to fibrotic changes in the lung. Abscess in any organ or tissue was classified as "Abscess" in the table without notation of its site. Other inflammatory changes, "Others" in the table, include all inflammatory lesions other than pneumonia, abscess, and colitis, such as chronic pyelonephritis, purulent pharyngitis, etc. Amyloid deposition was con-
PATHOBIOLOGYFEATLrRESOF THE SAM MODEL
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TABLE I. AUTOPSY FINDINGS AFTER NATURAL DEATH OF S A M M~CE (%)
Inflammation Strains
SAME1 SAMP2 SAMP3 SAMP6 SAMP7 SAMP8 SAMP9 SAMP10 SAMPI 1
Gen@
67-87 70-87 65-84 60-82 65-83 69-84 61-74 66-76 64-83
SAMP SAMR1 SAMR4 SAMR5 SAMR
59-72 60--73 57-72
No. of mice
Pneumonia
Ab
Col
Others
Amy
CK
Deg Tub
lnv
AN
Br
In
62 (29#) 76 (39) 66 (40) 49 (18) 88 (55) 46 (27) 35 (12) 102 (34) 41 (20)
19.4 14.5 30.3 10.2 5.7 10.9 20.0 15.7 31.7
8.1 1.3 1.5 6.1 2.3 2.2 0 1.0 2.4
35.5 23.7 27.3 32.7 27.3 26.1 42.9 20.6 29.3
0 2.6 1.5 8.2 0 4.3 0 3.9 4,9
3.2 9.2 18.2 6.1 5.7 13.0 2.9 7.8 4.9
82.0 85.5* 45.5 79.6 63.6 39.1 82.9 93.1" 58.5
46.8 63.2* 16.7 38.8 18.2 13.0 34.3 68.6* 46.3
6.5 14.5 4.5 14.3 4.5 8.7 8.6 30.4 7.3
0 2.6 0 0 0 4.3 0 0 0
6.5 0 1.5 2.0 0 0 0 9.8 4.9
565 (274)
16.6
2.7
28.0
2.7
8.1
71.9
40.7
12.4
0.7
3.2
77 (39) 52 (23) 75 (39)
16.9 25.0 14.7
1.3 3.8 0
31.2 34.6 22.7
1.3+ 11.5 33.3+
13.0 13.5 21.3
33.8* 32.5* 30.7*
5.2* 8.6* 12.0"
10.4 11.5 2.7
0 0 0
3.0 0 0
204 (101)
18.1
1.5
28.9
15.7
16.2
30.4
8.8
7.8
0
1.0
a
b
a
a
p-value: SAMP vs. SAMR**
Neoplasms Ang Strains
SAMP1 SAMP2 SAMP3 SAMP6 SAMP7 SAMP8 SAMP9 SAMPI0 SAMPI 1
Lymphoid cell neoplasms TLB
NTLB
IB
FCC
Unc
Combined
1.6 0 1.5 0 2.3 4.3 0 2.9 0
0 0 0 0 1.1 0 0 0 0
0 0 0 0 0 2.2 0 0 0
6.5 3.9 7.6 0 60.2* 32.6 42.9* 3.9* 0
1.6 0 0 0 0 0 0 7.8 0
4.8 2.6 6.1 0 55.7+ 23.9 42.9+ 0 0
0 1.3 0 0 1.1 2.2 0 1.0 0
SAMP
1.6
14.9
0.7
1.6
0.2
0.2
SAMRI SAMR4 SAMR5
0 0 0
1.3 1.9 0
1.3 1.9 1.4
14.3 15.4 1.4+
1.3 1.9 0
0 1.9 0
SAMR
0
1.0
1.5
9.8
1.0
0.5
p-value: SAMP vs. SAMR**
a
HS
a
GL
Others Malig
Ben
0 0 2.5 0 1.8 0 0 2.9 0
3.2 1.3 3.0 2.0 1.1 2.2 5.7 2.9 0
1.6 0 0 0 1.1 0 0 0 0
1.6 0 0 0 1.1 0 0 1.0 0
17.5
0.4
0.5
1.1
0.5
2.3
18.2 23.1 2.7+
19.5 15.4 9.5
3.9 1.9 0
3.9 3.8 4.1
46.2 + 13.0 12.8
0 1.9 8.0
13.7
14.7
2.0
3.9
12.7
3.4
a
4.8 0 0 0 1.1 0 2.9 1.0 0
CyOv
a
@generation, # number of females, *significantly different from other SAM strains by chi-square test, +significantly different from other SAMP or SAMR strains by chi-square test, **a: p < 0.001; b: p < 0.01, Br: bronchopneumonia; In: interstitial pneumonia; Ab: abscess; Col: colitis; Amy: amyloidosis; CK: contracted kidney; Deg Tub: degeneratio of renal tubule; Inv: invagination; AN: angionecrosis; Ang: angiitis; TLB: thymic lymphoblastic lymphoma; NTLB: nonthymic lymphoblastic lymphoma; IB: immunoblastic lymphoma; FCC: follicular center cell lymphoma; Unc: unclassifiable lymphoma; HS: histiocyfic sarcoma; GL: granulocytic leukemia; Malig: malignant neoplasms; Cy Ov: ovarian cyst; Ben: benign neoplasms other than ovarian cyst.
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Y. TAKEDA et a l
firmed in Congo red stained sections. Green birefringence of the Congo red strain under the polarizing microscope was considered to present amyloid (Takeshita et al., 1982). AA amyloid protein (secondary amyloidosis) was often observed in SAM mice with inflammatory lesions, particularly in SAMP2 and SAMP6. Senile amyloidosis, with apoA-1I protein deposits in organs or tissues, was observed more frequently and severely in SAMP1, -P2, -P7, -P9, -P10, and -P11 than in other strains of SAM. Contracted kidneys had a granular surface and were reduced in size according to the severity of the lesions. Destroyed nephrons were replaced by a proliferation of fibrous tissue. Degenerated renal tubules, "Deg Tub" in the table, showed vacuolar degeneration in their proximal convoluted sections. The vacuoles in the tubules did not show positive fat staining. Angionecrotic lesions were observed in the alimentary tract, pancreas, uterine adnexae, testes, ovaries, and kidneys. Angiitis was observed in the tongue, alimentary tract, pancreas, kidneys, uterine adnexae, and testes. Lymphoid cell neoplasms, histiocytic sarcoma, and granulocytic leukemia were diagnosed from gross and histological findings of the spleen, thymus, lymph nodes, bone marrow, and other affected organs, according to the classification of Pattengale and Taylor (1981, 1983). Three types of pathology in lymphoid cell neoplasms were observed in the SAM model: lymphoblastic lymphoma, immunoblastic lymphoma, and follicular center cell lymphoma. In this study, lymphoblastic lymphoma was classified into two types: thymic lymphoblastic lymphoma with definite neoplasm of thymic tissue in the anterior mediastinum, and nonthymic lymphoblastic lymphoma without any definite thymic neoplasm. Two lymphoid cell-derived neoplasms observed in a SAMP8 and a SAMR4 mouse are listed as " U n c " (unclassifiable) because of inadequate fixation (Table 1). Among the findings of routine postmortem examinations, amyloidosis, contracted kidney, immunoblastic lymphoma, histiocytic sarcoma, and ovarian cyst are age-associated pathologies the incidence of which increases with advancing age. The incidence of each pathologic finding was compared between SAMP and SAMR, among SAM strains, and also among SAMP or SAMR strains. The chi-square test on a two-by-two contingency table was used to test the statistical significance of differences between each genotype and all animals in the study as a ratio of animals with a pathology to those without incidence of certain common pathologic findings. As shown in Table 1, the incidence of amyloidosis (p < 0.001), contracted kidney (p < 0.001), and thymic lymphoblastic lymphoma (p < 0.001) was significantly higher in SAMP than in SAMR. On the other hand, colitis (p < 0.001), inflammatory changes other than pneumonia, abscess and colitis (p < 0.01), immunoblastic lymphoma (p < 0.001), histiocytic sarcoma (p < 0.001) and ovarian cyst (p < 0.001) were more frequent in SAMR than in SAMP. The incidence of bronchopneumonia and abscess did nol differ significantly between SAMP and SAMR. When the incidence of each abnormality was compared among all the SAM strains, that of amyloidosis and contracted kidney was significantly greater in SAMP2 and SAMP10, and was significantly lower in SAMR1, SAMR4, and SAMR5, than in the other genotypes of SAM. SAMP7 and SAMP9 had a significantly higher incidence of lymphoid cell neoplasms than did the others. A significantly lower incidence of lymphoid cell neoplasms was observed in SAMP10 than in the other strains of SAM. In the statistical analysis by the chi-square test of all the SAM strains, a p-value less than 0.001 was considered to be statistically significant. Among SAMP strains, SAMP7 (p < 0.001) and SAMP9 (p < 0.001) had a higher incidence of thymic lymphoblastic lymphoma than did the other genotypes. Among the SAMR strains, SAMR5 had a significantly lower incidence of immunoblastic lymphoma (p < 0.01) and lymphoid cell neoplasms (p < 0.01) than did the others. The incidence of ovarian cysts was significantly greater (p < 0.01) in SAMR1 than in the others. Table 2 shows a summary of the three most frequently seen autopsy findings in each SAM
121
PATHOBIOLOGYFEATURESOF THE SAM MODEL
strain. In all the SAMP strains, amyloidosis was the most common autopsy finding with an incidence of 39.1-93.1% and was highest in SAMP10. The incidence in SAMR was 30.733.8%. Five of the SAMP strains, SAMP1, -P2, -P6, -P10, and -P11, had contracted kidney as the second most frequent autopsy finding; and in three, SAMP7, -P8, and -P9, it was lymphoid cell neoplasm. The third most common autopsy finding in seven strains of SAMP was abscess. In SAMP10 it was Deg Tub, degeneration of the renal tubules. Ovarian cyst (46.2%) and colitis (33.3%) were the most common pathologies in SAMR1 and SAMR5, respectively. The pathologic changes causing death were assessed in 1337 SAMP and 453 SAMR mice, in some by gross observation only. These included pneumonia, abscess, colitis, amyloidosis, contracted kidney, invagination, neoplasms, including lymphoid cell neoplasms, histiocytic sarcoma and other neoplasms, miscellaneous pathologic findings other than the above, and unknown causes. In the statistical analysis of the causes of death, the chi-square test was used. As shown in Table 3, colitis (p < 0.001), histiocytic sarcoma (p < 0.001), and unknown causes (p < 0.01) were more frequent causes of death in SAMR than in SAMP, and contracted kidney (p < 0.001) and lymphoid cell neoplasms (p < 0.001) were significantly more frequent in SAMP than in SAMR. In a comparison of the causes of death among all the SAM strains at the significance level of p < 0.001, pneumonia was a more frequent cause of death in SAMP3 and SAMR1 than in the other strains. In SAMP1 and SAMP10, contracted kidney was a more frequent cause of death than in the other genotypes. SAMP7, -P8, and -P9 mice died of lymphoid cell neoplasms significantly more frequently than did other strains of SAM. SAMR5 mice died more frequently without overt pathologic changes than did other strains of SAM. On the other hand, abscess in SAMP10, contracted kidney in SAMP7, -P8, -R1, and -R4, lymphoid cell neoplasms in SAMP2, -P6, -P10, and -P11, and unknown causes in SAMP7 and -P10 were less frequent causes of death than in the other strains of SAM. TABLE 2. TOP THREE AUTOPSYFINDINGS IN SAM STRAINS Strains
SAMP SAMP 1 SAMP2 SAMP3 SAMP6 SAMP7 SAMP8 SAMP9
1
Amy Amy Amy Amy
2
3
CK CK Br CK LN LN LN CK CK
Ab Ab Ab Ab Ab Ab Ab
Amy, Cy Ov
Ab
Ab Co
Amy Amy
HS Br Ab
Amy
Amy Amy
SAMP10
Amy
SAMPI 1 SAMR SAMR1 SAMR4 SAMR5
Amy
Deg Tub Br
Amy: amyloidosis; CK: contracted kidney; Ab: abscess; Br: bronchopneumonia; LN: lymphoid cell neoplasms; Deg Tub: degeneration of renal tubule; HS: histiocytic sarcoma; Cy Ov: ovarian cyst; Co: colitis.
T. TAKEDA et al.
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TABLE 3. AUTOPSY FINDINGSCAUSING DEATH(%)
Strains
Gen+
No.
Inflammation
Amy
CK
lnv
Neoplasms
Others
Unk
of mice
SAMPI SAMP2 SAMP3 SAMP6 SAMP7 SAMP8 SAMP9 SAMPI0 SAMPI1
SAMR
Ab
112 200 97 195 247 132 47 204 103
11.6 24.0 34.0* 15.9 22.7 15.2 6.4 1(I.8 24.3
22.3 23.5 20.6 20.0 16.6 18.2 19.2 6.8 13.6
0.9 3.5 0 2.1 0 4,5 0 1.0 0
0.9 0 0 2.6 0 0 0 0.5 2.9
28.6 21.5 3.1 17.4 7.7* (/.8" 8.5 47.5* 22.3
1337
17.2
t7.4
1.5
0.7
19.1
237 139 77
29.1" 23.0 16.9
17.7 15.8 10.4
2.5 14.4 1.3
0.8 0 1.3
453
25.2
15.9
6.0
0.7
67-8l 70-86 64-82 66-80 65-81 69-81 61-70 66-73 64-78
SAMP SAMRI SAMR4 SAMR5
Pn
59-73 60-71 55-70
p-value: SAMP vs. SAMR**
a
Col
a
LN
HS
ON
1.8 3.0 0 0 0 6.0 0 1,0 0
2.7 1.5" 6.2 0* 38.5* 23.5* 46.8* 1.5" 0*
0.9 0 0 0 0.4 0 0 0 0
1.8 0 0 0 0.8 0 0 0.5 0
11.6 9.5 12.4 14.3 3.6 8.3 8.5 20.1 8.7
17.0 23.5 23.7 27.7 9.7* 23.5 10.6 10.3" 28.2
1.3
12.2
0.1
0.4
10.9
18.9
3.0* 2.2* 1.3
t.3 0.7 0
5.5 10. t 2.6
4.6 6.5 5.2
3.0 2,1 2.6
9.7 5.0 16,9
22.8 20.1 41.5"
2.4
0.9
6.4
5.3
2,6
9,5
a
b
a
28.1
+generation; *significantly different from other SAM strains by ehi-square test; **a: p < 0.001, b: p < 0.01; Pn: pneumonia; Ab: abscess; Col: colitis; Amy: amyloidosis; CK: contracted kidney; lnv: invagination; LN: lymphoid cell neoplasms; HS: histocytic sarcoma; ON: other neoplasms; Unk: unknown.
The top three causes of death in each SAM strain are summarized in Table 4. Lymphoid cell neoplasm was the most common cause of death in SAMP7, -P8, and -P9; while contracted kidney was the most frequent cause of death in SAMP1 (28.6%) and SAMP10 (47.5%). The cause of death was unknown in 41.5% of SAMR5, in 28.2% of SAMP11, and in 27.7% of SAMP6. In these three, no known causes of death were of higher incidence. SAMR5, a strain with normal aging, had fewer fatal pathologies such as lymphoid cell neoplasms and histiocytic sarcoma, contracted kidney, or inflammatory changes (except for colitis) than did the other strains (Table 1). Interestingly enough, amyloidosis with the highest or second highest incidence in autopsy findings in all SAM strains, was only a minor cause of death (Tables 1-4), because other more severe pathologies that often coexisted with amyloidosis, such as malignant neoplasms, contracted kidney, etc., were the causes of death. PATHOBIOLOGICAL FEATURES REVEALED BY SYSTEMATICALLY DESIGNED STUDIES Besides the routine postmortem examinations, several studies were conducted to assess systematically and dynamically various age-related pathologic changes. A series of behavioral studies on SAMP8 and SAMP10 disclosed age-related deficits in learning and memory (Miyamoto et al., 1986; Yagi et al., 1988; Ohta et al., 1989; Flood and Morley, 1992, 1993; Shimada et al., 1993). In SAMP10, age-related spontaneous brain atrophy was also observed. Miyamoto et al. (1992) reported a characteristic emotional disorder, reduced anxiety-like behavior, and a profound disorder of circadian rhythm, in addition to the deficits in learning and memory, in SAMP8/Ta reared under specific pathogen-free (SPF) conditions at Takeda Chemical Industries, Ltd. To clarify the pathogenic mechanisms in the learning and memory deficits
PATHOBIOLOGY FEATURES OF THE SAM MODEL
123
TABLE4. TOPTHREECAUSESOFDEATHIN SAM STRAINS Strains
SAMP SAMP1 SAMP2 SAMP3 SAMP6 SAMP7 SAMP8 SAMP9 SAMP10 SAMP11 SAMR SAMRI SAMR4 SAMR5
1
2
3
CK Pn Pn Unk LN LN LN CK Unk
Ab Ab Unk Ab Pn Unk Ab Others Pn
Unk Unk Ab CK Ab Ab Unk Pn CK
Pn Pn Unk
Unk Unk Pn
Ab Ab Others
CK: contractedkidney;Ab: abscess;Unk:unknown;Pn: pneumonia, including bronchopneumoniaand interstitial pneumonia; LN: lymphoidcell neoplasms; Others: see text. in SAMP8 and SAMP10, a series of neuropathological, neurochemical, and neuropharmacological experiments were performed (Shimada et al., 1992; Nomura et al., 1994; Flood and Morley, 1994). In the course of development of SAM, we became aware of spontaneous leg fractures in a few aged SAM mice. Therefore, we began to examine systematically the age-related changes in bone in several strains of SAM and found that SAMP6 mice showed senile osteoporosis characterized by a low peak bone mass (Matsushita et al., 1986). A series of studies to clarify the pathogenesis of osteoporosis in SAMP6 is now under way in our laboratory (Tsuboyama et al., 1989a,b, 1993; Takahashi et al., 1994; Okamoto et al., 1995) as well as in others (Kawase et al., 1989; Suda et al., 1994). A systematic study on the condyle of the temporomandibular joint in several strains of SAM revealed that all strains of SAM develop degenerative joint disease. However, the short-lived SAMP strains develop degenerative changes earlier than do the SAM strains. SAMP3 mice are the first to show degenerative joint changes (approximately 50% at seven to nine months of age and 100% over 12 months of age) and thereafter have the highest incidence of severe changes with overt deformity (Chen et al., 1989). Examinations of the eyes and ears showed age-related cataracts in SAMP9 and age-related hearing impairment in SAMP1 and SAMR1. Persistence of the hyaloid vascular system is a necessary, but not the sole, factor in SAMP9 cataracts (Hosokawa et al., 1988, 1993). The persistence of the hyaloid vascular system might affect the breakdown of the lens capsule, a trigger of cataract formation, and subsequently lead to increased Ca content and transglutaminase activity in the lens to reduce the solubility of [~-crystallins (Ashida et al., 1994). SAMP1 and SAMR1 showed an age-related auditory loss expressed as elevated thresholds, prolongation of interpeak intervals I-III and 1-IV, and decreased amplitude of wave I assessed by auditory brainstem responses (ABRs) (Saitoh et al., 1994). SAMP1 showed a more rapid and
124
T. TAKEDAet al
severe auditory loss with age than did SAMRI. Morphological studies showed an age-related decrease in cell density as well as the size of spiral ganglion neurons in SAMPI and SAMR1 (Saitoh et al., 1994). An age-related loss of the inner and outer hair cells was observed in both SAMP1 and SAMRI. The changes in the spiral ganglion neurons and hair cells, however, appeared earlier and progressed more rapidly in SAMP1 than in SAMR1. Furthermore, SAMPI showed greater age-related atrophy of the stria vascularis than did SAMR1. These results suggest that hearing impairment in SAM is due to a combination of sensory and strial presbycusis as well as to neural presbycusis (Saitoh et al., 1995). Morphometric and physiological studies of the respiratory system showed senile hyperinflation of the lungs in SAMP1 and SAMR 1. Histologic observation, however, revealed no evidence of destruction of the alveolar wall or elastic fibers in the lungs. The hyperinflationary changes occurred similarly in both SAMPI and SAMR1, but in an accelerated manner in SAMPI ; and, finally, senile hyperinflation of the lungs occurred in the senescence phase in both strains (Kurozumi et al., 1994). A series of immunobiological studies was performed. In vivo experiments showed that primary antibody response and delayed-type hypersensitivity (DTH) reaction to sheep red blood cells (SRBC) were significantly more impaired by six months of age in SAMP1 and SAMP2 than in SAMR1 and ordinary strains such as C3H/He and B10.BR (Mitsuoka and Hanada, 1988). To clarify the nature of the immunobiological defects observed in vivo, in vitro experiments were conducted, and it became evident that SAMP1 mice had a profound defect in antibody response to a T-dependent antigen, SRBC as early as two months of age and a negligible response at a later age (Hosokawa et al., 1987a). This impaired antibody response was considered to be closely related to dysfunction of T-helper cells, while other T-cell activities such as mixed lymphocyte reaction, cytotoxic T lymphocyte generation and DTH reaction remained normal. B-cell and antigen-presenting cell activities were also normal (Hosokawa et al., 1987b). Recently, Abe e t al. (1994) reported that a helper T-cell defect similar to that in SAMP1 was found in SAMP8 mice that show characteristic deficits in learning and memory with advancing age. Furthermore, low natural killer (NK) cell activity was observed in SAMP8. Among the pathobiological findings in SAM revealed by routine postmortem examinations and also by systematic studies, there are pathologic phenotypes that are often characteristic enough to distinguish among all the SAM strains, as shown in Table 5. Furthermore, most of the "age-dependent" geriatric disorders seen in humans, which were described as "inevitable disorders as a direct consequence of physiological senescence" by Cotran e t al. (1989), are included in the pathobiological phenotypes in SAM: these are osteoporosis, degenerative joint disease, cataract, hyperinflation of lungs, and hearing impairment. The evidence provides support for our proposal that the SAM model is a valid one for research on aging. CONCLUSIONS In general, one of the most important issues in the development and maintenance of animal models for biomedical research is the maintenance of the expression of the characteristic phenotypes. The SAM model, the characteristics of which have been strictly monitored pathobiologically as well as genetically (Festing, 1992, 1993), is expected to clarify the mechanisms of age-associated pathologies and to provide effective methods of modulating or ameliorating the advance of senescence and the disease processes involved in age-associated pathologies. The fundamental mechanisms of accelerated, as well as primary, aging processes can be demonstrated in the SAM model.
PATHOBIOLOGYFEATURESOFTHE SAMMODEL
125
TABLE 5. PATHOBIOLOGICALPHENOTYPESIN SAM MICE Strains SAMP SAMP1 SAMP2 SAMP3 SAMP6 SAMP7 SAMP8 SAMP9 SAMP10 SAMP11 SAMR SAMR1 SAMR4 SAMR5
Phenotypes
Senile amyloidosis, contracted kidney, impaired immune response, hyperinflation of lungs, hearing impairment. Senile and secondary amyloidosis, contracted kidney, impaired immune response. Degenerative temporomandibular joint disease. Senile osteoporosis, secondary amyloidosis. Thymic lymphoblastic lymphoma, senile amyloidosis. Deficits in learning and memory, emotional disorder (reduced anxiety-like behavior),* impaired immune response. Cataract, thymic lymphoblastic lymphoma, senile amyloidosis. Brain atrophy, deficits in learning and memory, emotional disorder (depressive behavior),* contracted kidney, senile amyloidosis. Contracted kidney, senile amyloidosis. Nonthymic lymphoma,t histiocytic sarcoma, ovarian cyst. Nonthymic lymphoma,t histiocytic sarcoma. Colitis
*Reported in SAMP8/'ra or SAMP10/Ta mice reared under SPF conditions at Takeda Chemical Industries Ltd. tlncludes nonthymic lymphoblastic lymphoma, immunoblastic lymphoma and follicular center cell lymphoma. Acknowledgments--This work was supported in part by grants from the Ministry of Education, Science and Culture and the Ministry of Health and Welfare, Japan. Gratitude is extended to Drs. W. H. Chen, A. Ohta, and N. Seriu, and K. Kadota and K. Yasuoka for technical assistance. We are also grateful to Dr. A. Cary for critical reading of the manuscript. REFERENCES ABE, Y., YUASA, M., KAJIWARA, Y., and HOSONO, M. Deficits of immune cells in the Senescence-Accelerated Mouse: A model for learning and memory deficits in the aged. Cell Immunol. 157, 59-69, 1994. ASHIDA, Y., TAKEDA, T., and HOSOKAWA, M. Protein alterations in age-related cataract associated with a persistent hyaloid vascular system in senescence-accelerated mouse (SAM). Exp. Eye Res. 59, 467-474, 1994. CHEN, W.H., HOSOKAWA, M., TSUBOYAMA, T., ONO, T., IIZUKA, T., and TAKEDA, T. Age-related changes in the temporomandibular joint of the Senescence Accelerated Mouse (SAM): SAM-P/3 as a new murine model of degenerative joint disease. Am. J. Pathol. 135, 379-385, 1989. COTRAN, R.S., KUMAR, Y., and ROBBINS, S.L. Disease of aging. In: Robbins Pathologic Basis of Disease, pp. 543-551, W.B. Saunders Company, Philadelphia, PA, 1989. FESTING, M.F.W. Origins and characteristics of inbred strains of mice. Mouse Genome 90, 236-237, 1992. FESTING, M.F.W. Origins and characteristics of inbred strains of mice. Mouse Genorne 91, 471-472, 1993. FLOOD, J.F. and MORLEY, J.E. Early onset of age-related impairment of aversive and appetitive learning in the SAM-P/8 mouse. J. Gerontol. 47, B52-B59, 1992. FLOOD, J.F. and MORLEY, J.E. Age-related changes in foot shock avoidance acquisition and retention in senescence accelerated mouse (SAM). Neurobiol. Aging 14, 153-157, 1993. FLOOD, J.F., and MORLEY, J.E. Age-related changes in the pharmacological improvement of retention in SAM8 mice. In: The SAM Model of Senescence, Takeda, T. (Editor), pp. 89-94, Excerpta Medica, Elsevier Science B.V., Amsterdam, 1994. HIGUCHI, K., MATSUMURA, A., HONMA, A., TAKESHITA, S., HASHIMOTO, K., HOSOKAWA, M., YASUHIRA, K., and TAKEDA, T. Systemic senile amyloid in Senescence-Accelerated Mice: A unique fibril protein demonstrated in tissues from various organs by the unlabeled immunoperoxidase method. Lab. Invest. 48, 231-240, 1983. HIGUCHI, K., NAIKI, H., KITAGAWA, K., HOSOKAWA, M., and TAKEDA, T. Mouse senile amyloidosis: ASsAM
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amyloidosis in mice is universally present as a systemic age-associated amyloidosis. Virchows Arch. B Cell Pathol. 60, 231-239, 1991a. HIGUCH1, K., KITAGAWA, K., NAIKI, H., HANADA, K., HOSOKAWA, M., and TAKEDA, T. Polymorphism of apolipoprotein A-II (apoA-II) among inbred strains of mice. Relationship between the molecular type of apoA-II and mouse senile amyloidosis. Biochem. J. 279, 427-433, 1991b. HOSOKAWA, M., ASHIDA, Y., TSUBOYAMA, T., CHEN, W.H., and TAKEDA, T. Cataract in Senescence Accelerated Mouse (SAM) 2. Development of a new strain of mouse with late-appearing cataract. Exp. Eye Res. 47, 629-640, 1988. HOSOKAWA, M., ASHIDA, Y., MATSUSHITA, T., TAKAHASHI, K., and TAKEDA, T. Persistent hyaloid vascular system in age-related cataract in a SAM strain of mouse. Eap. Eye Res. 57, 427-434, 1993. HOSOKAWA, T., HOSONO, M., HIGUCHI, K., AOIKE, A., KAWAI, K., and TAKEDA, T. Immune responses in newly developed short-lived SAM mice 1. Age-associated early decline in immune activities of cultured spleen cells. Immunology 62, 419-423, 1987a. HOSOKAWA, T., HOSONO, M., HANADA, K., AOIKE, A., KAWAI, K., and TAKEDA, T. Immune responses in newly developed short-lived SAM mice II. Selectively impaired T-helper cell activity in in vitro antibody response. Immunology 62, 425-429, 1987b. KAWASE, M., TSUDA, M., and MATSUO, T. Accelerated bone resorption in senescence-accelerated mouse (SAMP/6). J. Bone Miner. Res. 4, 359-364, 1989. KUROZUMI, M., MATSUSHITA, T., HOSOKAWA, M., and TAKEDA, T. Age-related changes in lung structure and function in the Senescence-Accelerated Mouse (SAM): SAM-P/I as a new murine model of senile hyperinflation of lung. Am. J. Respir. Crit. Care Med. 149, 776-782, 1994. MATSUSHITA, M., TSUBOYAMA, T., KASAI, R., OKUMURA, H., YAMAMURO, T., HIGUCHI, K., HIGUCHI, K., KOHNO, A., YONEZU, T., UTANI, A., UMEZAWA, M., and TAKEDA, T. Age-related changes in bone mass in the Senescence Accelerated Mouse (SAM): SAM-R/3 and SAM-P/6 as new murine models for senile osteoporosis. Am. J. Pathol. 125, 276-283, 1986. MITSUOKA, A, and HANADA, K. In vivo immune reactivities in Senescence Accelerated Mice (SAM). In: Proceedings of The First SAM Kyoto Symposium, Takeda, T., Matsuo, T., Akiguchi, I., and Hosokawa, M. (Editors), pp. 11-20, Fuji Printing Business Company, Kyoto, Japan, 1988. MIYAMOTO, M., KIYOTA, Y., YAMAZAKI, N., NAGAOKA, A., MATSUO, T., NAGAWA, Y., and TAKEDA, T. Age-related changes in learning and memory in the senescence-accelerated mouse (SAM). Physiol. Behav. 38, 399-406, 1986. MIYAMOTO, M., KIYOTA, Y., NISHIYAMA, M., and NAGAOKA, A. Senescence- accelerated mouse (SAM): Age-related reduced anxiety-like behavior in the SAM-P/8 strain. Physiol. Behav. 51,979-985, 1992. NOMURA, Y., KITAMURA, Y., ZHAO, X.H., OHNUKI, T., TAKEI, M., YAMANAKA, Y., KITAYAMA, Y., and NISHIYA. T. Neurochemical studies on aging in SAM brain. In: The SAM Model of Senescence, Takeda, T. (Editor), pp. 83-88, Excerpta Medica, Elsevier Press B.V., Amsterdam, 1994. OHTA, A., HIRANO, T., YAGI, H., TANAKA, S., HOSOKAWA, M., and TAKEDA, T. Behavioral characteristics of the SAM-P/8 strain in Sidman active avoidance task. Brain Res. 498, 195-198, 1989. OKAMOTO, Y., TAKAHASH1, K., TORIYAMA, K., TAKEDA, N., KITAGAWA, K., HOSOKAWA, M., and TAKEDA, T. Femoral peak bone mass and osteoclast number in an animal model of age-related spontaneous osteopenia. Anat. Rec. 242, 21-28, 1995. PATTENGALE, P.K. and TAYLOR, C.R. Immunomorphologic classification of murine lymphomas and related leukemias. In: Proceedings of Rodent Lymphoma Workshop, March 4-5, 1981, Jefferson, AR, pp. 22-23, National Center Toxicol. Res. Press, Jefferson, AR, 1981. PATTENGALE, P.K. and TAYLOR, C.R. Experimental model of lymphoproliferative disease: The mouse as a model for human nonHodgkin's lymphoma and related leukemias. Am. J. Pathol. 113, 237-265, 1983. SAITOH, Y., HOSOKAWA, M., SHIMADA, A., WATANABE, Y., YASUDA, N., TAKEDA, T., and MURAKAMI, Y. Age-related hearing impairment in senescence-accelerated mouse (SAM). Hear. Res. 75, 27-37, 1994. SAITOH, Y., HOSOKAWA, M., SHIMADA, A., WATANABE, Y., YASUDA, N., MURAKAMI, Y., and TAKEDA, T. Age-related cochlear degeneration in senescence-accelerated mouse. Neurobiol. Aging 16, 129-136, 1995. SHIMADA, A., OHTA, A., AKIGUCHI, I., and TAKEDA, T. Inbred SAM-P/10 as a mouse model of spontaneous inherited brain atrophy. J. Neuropathol. Exp. Neurol. 51, 440-450, 1992. SHIMADA, A., OHTA, A., AKIGUCHI, I., and TAKEDA, T. Age-related deterioration in conditional avoidance task in the SAM-P/10 mouse, an animal model of spontaneous brain atrophy. Brain Res. 608, 266-272, 1993. SUDA, T., MIYAMA, K., UCHIYAMA, Y., KATAGIRI, T., YAMAGUCHI, A., and SATO, T. Osteoporotic bone
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changes in SAMP6 are due to a decrease in osteoblast progenitor cells. In: The SAM Model of Senescence, Takeda, T. (Editor), pp. 47-52, Excerpta Medica, Elsevier Press B.V., Amsterdam, 1994. TAKAHASHI, K., TSUBOYAMA, T., MATSUSHITA, M., KASAI,R., OKUMURA, H., YAMAMURO, T~, OKAMOTO, Y., TORIYAMA, K., KITAGAWA, K., and TAKEDA, T. Modification of strain-specific femoral bone density by bone marrow derived factors administered neonatally: A study on the spontaneously osteoporotic mouse, SAMP6. Bone Miner. 24, 245-255, 1994. TAKEDA, T., HOSOKAWA, M., TAKESHITA, S., IRINO, M., HIGUCHI, K., MATSUSHITA, T., TOMITA, Y., YASUHIRA, K., HAMAMOTO, H., SHIMIZU, K., ISHII, M., and YAMAMURO, T. A new murine model of accelerated senescence. Mech. Ageing Dev. 17, 183-194, 1981. TAKEDA, T., HOSOKAWA, M., and HIGUCHI, K. Senescence-Accelerated Mouse (SAM): A novel murine model of accelerated senescence. J. Am. Geriatr. Soc. 39, 911-919, 1991. TAKEDA, T., HOSOKAWA, M., HIGUCHI, K., HOSONO, M., AKIGUCHI, I., and TAKEDA, T. A novel murine model of aging, Senescence-Accelerated Mouse (SAM). Arch. Gerontol. Geriatr. 19, 185-192, 1994. TAKESHITA, S., HOSOKAWA, M., IRINO, M., HIGUCHI, K., SHIMIZU, K., YASUHIRA, K., and TAKEDA, T. Spontaneous age-associated amyloidosis in Senescence-Accelerated Mouse (SAM). Mech. Ageing Dev. 20, 13-23, 1982. TSUBOYAMA, T., TAKAHASHI, K., MATSUSHITA, M., OKUMURA, H., YAMAMURO, T., UMEZAWA, M., and TAKEDA, T. Decreased endosteal formation during cortical bone modelling in SAM-P/6 mice with a low peak bone mass. Bone Miner. 7, 1-12, 1989a. TSUBOYAMA, T., MATSUSHITA, M., OKUMURA, H., YAMAMURO, T., HANADA, K., and TAKEDA, T. Modification of strain-specific femoral bone density by bone marrow chimerism in mice: A study on spontaneously osteoporotic mouse (SAM-P/6). Bone 10, 269-277, 1989b. TSUBOYAMA, T., TAKAHASHI, K., YAMAMURO, T., HOSOKAWA, M., and TAKEDA, T. Cross mating study on bone mass in the spontaneously osteoporotic mouse (SAM-P/6). Bone Miner. 23, 57--64, 1993. YAGI, H., KATOH, S., AKIGUCHI, I., and TAKEDA, T. Age-related deterioration of ability of acquisition in memory and learning in Senescence Accelerated Mouse: SAM-P/8 as an animal model of disturbances in recent memory. Brain Res. 474, 86--93, 1988.
PII S0531-5565(96)00068-X
PATHOBIOLOGY OF THE SENESCENCE-ACCELERATED MOUSE (SAM)
TOSHIO TAKEDA,l TAKATOSHIMATSUSHITA,2 MAFUMIKUROZUMI,3 KIMINOBUTAKEMURA,2 KEIICHI HIGUCHI2 and MASANORIHOSOKAWA2 IThe Council for SAM Research, Departmentof Senescence Biology and 2Departmentsof Senescence Biology and 3ClinicalLaboratory, Chest Disease Research Institute,Kyoto University,Sakyo-ku,Kyoto606, Japan
Abstract--Routine postmortem examinations and the pathobiological features revealed by systematically designed studies have shown several pathologic phenotypes that are often characteristic enough to differentiate among the various SAM strains: senile amyloidosis in SAMP1, -P2, -P7, -P9, -P10, and -P11; secondary amyloidosis in SAMP2 and -P6; contracted kidney in SAMP1, -P2, -P10, and - P l l ; immunoblastic lymphoma in SAMR1 and -R4; histiocytic sarcoma in SAMR1 and -R4; ovarian cysts in SAMR1; impaired immune response in SAMP1, -P2, and -P8; hyperinflation of the lungs in SAMP1; hearing impairment in SAMPI; degenerative temporomandibular joint disease in SAMP3; senile osteoporosis in SAMP6; deficits in learning and memory, in SAMP8 and -P10; emotional disorders in SAMP8 and -P10; cataracts in SAMP9; and brain atroFhy in SAMP10. These are all age-associated pathologies, the .incidence and severity of which increase with advancing age. The SAM model in which these pathobiological features have been carefully monitored will be a valuable tool in the clarification of the pathogenic mechanisms of age-associated pathologies and in the research for effective methods to modnLate or ameliorate these pathologies. Copyright © 1997 Elsevier Science Inc.
Key Words: Senescence-Accelerated Mouse, SAM, aging, senescence, pathobioiogy, pathologic phenotypes, age-associated pathologies, autopsy findings, postmortem examination INTRODUCTION SENESCENCE-ACCELERATED Mouse (SAM) has been under development at Kyoto University since 1970 by selective sister-brother matings of mice of the AKR/J strain donated by the Jackson Laboratory (Bar Harbor, ME) in 1968. At present, there are 12 lines: senescence-prone inbred strains (SAMP) include S A M P I , SAMP2, SAMP3; SAMP6, SAMP7, SAMP8, SAMP9, SAMP10, and S A M P l l ; and senescence-resistant strains (SAMR) SAMR1, SAMR4, and SAMR5 (Takeda et al., 1981, 1991, 1994).
Correspondence to: Toshio Takeda, The Council for SAM Research, Department of Senescence Biology, Chest Disease Research Institute, Kyoto University,Sakyo-ku, Kyoto 606, Japan 117
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T. TAKEDA
e¢ al
In the first report on SAM (Takeda et al., 1981), amyloid deposition, abscess, thymic and nonthymic lymphoma, and pneumonia were cardinal autopsy findings in both the R and the P series, progenitors of SAMR and SAMP. The pathologic findings of recent routine postmortem examinations of SAMP and SAMR strains are described in this report, and the causes of death are analyzed. In addition, a series of extensive studies on the nervous, skeletal, sensory, respiratory, and immune systems have been conducted on living as well as dead SAM mice in a more dynamic approach to age-related pathobiological changes. Our findings include many pathobiological features, some of which are specific for each strain of the SAM model. AUTOPSY FINDINGS OF ROUTINE POSTMORTEM EXAMINATIONS Five hundred and sixty-five SAMP mice, 291 males and 274 females (62, 76, 66, 49, 88, 46, 35, 102, and 41 SAMP1, -P2, -P3, -P6, -P7, -P8, -P9, -P10, and -PI 1, respectively) and 204 SAMR mice, 103 males and 101 females (77, 52, and 75 SAMR1, -R4, and -R5, respectively), including both fresh carcasses and mice killed while moribund, were examined grossly and histologically. All these mice were reared under conventional conditions at the Chest Disease Research Institute, Kyoto University. The organs examined were the trachea, tongue, pharynx, larynx, salivary glands, thymus, thyroid, liver, kidney, spleen, lungs, heart, aorta, adrenal glands, abdominal skin, esophagus, alimentary tract, pancreas, and reproductive organs. As a rule, other organs and tissues, such as brain, spinal cord, nasal cavity, middle and inner ear, skeletal tissues, lymph nodes, etc., were not examined routinely. All organs and tissues were fixed in 10% buffered formalin and embedded in paraffin. The stains used were hematoxylin-eosin (4 Ia,m sections) and alkaline Congo red (6 ~m sections). When necessary, special stains were used: fat, PTAH (phosphotungstic acid hematoxylin), Azan-Mallory, and periodic acid-Schiff (PAS) or immunohistochemical stains with anti-apoA-II and anti-AA antisera for the identification of amyloid protein (Higuchi et al., 1983, 1991a,b). The gross and microscopic pathomorphological findings are summarized in Table 1, and the incidence of each pathologic finding is listed. The definite presence of any abnormality was considered positive, regardless of its severity. For example, the incidence of amyloidosis indicates the presence of amyloid definitely observed in any organ or tissue regardless of the severity of amyloid deposition. The autopsy findings were classified according to the type of change: inflammation, including bronchopneumonia, interstitial pneumonia, abscess, colitis, and other inflammatory changes; amyloidosis; contracted kidney; degeneration of renal tubules; invagination; angionecrosis; angiitis; lymphoid cell neoplasms, including thymic lymphoblastic lymphoma, nonthymic lymphoblastic lymphoma, immunoblastic lymphoma, follicular center cell lymphoma, and unclassifiable lymphomas; histiocytic sarcoma; granulocytic leukemia; and other neoplasms, including malignant neoplasms, ovarian cysts, and other benign neoplasms, such as hemangioma and pulmonary adenoma. Other miscellaneous pathologies were observed sporadically, such as pulmonary edema, hyperplasia of the extrahepatic bile duct, etc., but they are not listed. Interstitial pneumonia was often observed in mice with an abscess in any organ or tissue, but the lesions did not advance to fibrotic changes in the lung. Abscess in any organ or tissue was classified as "Abscess" in the table without notation of its site. Other inflammatory changes, "Others" in the table, include all inflammatory lesions other than pneumonia, abscess, and colitis, such as chronic pyelonephritis, purulent pharyngitis, etc. Amyloid deposition was con-
PATHOBIOLOGYFEATLrRESOF THE SAM MODEL
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TABLE I. AUTOPSY FINDINGS AFTER NATURAL DEATH OF S A M M~CE (%)
Inflammation Strains
SAME1 SAMP2 SAMP3 SAMP6 SAMP7 SAMP8 SAMP9 SAMP10 SAMPI 1
Gen@
67-87 70-87 65-84 60-82 65-83 69-84 61-74 66-76 64-83
SAMP SAMR1 SAMR4 SAMR5 SAMR
59-72 60--73 57-72
No. of mice
Pneumonia
Ab
Col
Others
Amy
CK
Deg Tub
lnv
AN
Br
In
62 (29#) 76 (39) 66 (40) 49 (18) 88 (55) 46 (27) 35 (12) 102 (34) 41 (20)
19.4 14.5 30.3 10.2 5.7 10.9 20.0 15.7 31.7
8.1 1.3 1.5 6.1 2.3 2.2 0 1.0 2.4
35.5 23.7 27.3 32.7 27.3 26.1 42.9 20.6 29.3
0 2.6 1.5 8.2 0 4.3 0 3.9 4,9
3.2 9.2 18.2 6.1 5.7 13.0 2.9 7.8 4.9
82.0 85.5* 45.5 79.6 63.6 39.1 82.9 93.1" 58.5
46.8 63.2* 16.7 38.8 18.2 13.0 34.3 68.6* 46.3
6.5 14.5 4.5 14.3 4.5 8.7 8.6 30.4 7.3
0 2.6 0 0 0 4.3 0 0 0
6.5 0 1.5 2.0 0 0 0 9.8 4.9
565 (274)
16.6
2.7
28.0
2.7
8.1
71.9
40.7
12.4
0.7
3.2
77 (39) 52 (23) 75 (39)
16.9 25.0 14.7
1.3 3.8 0
31.2 34.6 22.7
1.3+ 11.5 33.3+
13.0 13.5 21.3
33.8* 32.5* 30.7*
5.2* 8.6* 12.0"
10.4 11.5 2.7
0 0 0
3.0 0 0
204 (101)
18.1
1.5
28.9
15.7
16.2
30.4
8.8
7.8
0
1.0
a
b
a
a
p-value: SAMP vs. SAMR**
Neoplasms Ang Strains
SAMP1 SAMP2 SAMP3 SAMP6 SAMP7 SAMP8 SAMP9 SAMPI0 SAMPI 1
Lymphoid cell neoplasms TLB
NTLB
IB
FCC
Unc
Combined
1.6 0 1.5 0 2.3 4.3 0 2.9 0
0 0 0 0 1.1 0 0 0 0
0 0 0 0 0 2.2 0 0 0
6.5 3.9 7.6 0 60.2* 32.6 42.9* 3.9* 0
1.6 0 0 0 0 0 0 7.8 0
4.8 2.6 6.1 0 55.7+ 23.9 42.9+ 0 0
0 1.3 0 0 1.1 2.2 0 1.0 0
SAMP
1.6
14.9
0.7
1.6
0.2
0.2
SAMRI SAMR4 SAMR5
0 0 0
1.3 1.9 0
1.3 1.9 1.4
14.3 15.4 1.4+
1.3 1.9 0
0 1.9 0
SAMR
0
1.0
1.5
9.8
1.0
0.5
p-value: SAMP vs. SAMR**
a
HS
a
GL
Others Malig
Ben
0 0 2.5 0 1.8 0 0 2.9 0
3.2 1.3 3.0 2.0 1.1 2.2 5.7 2.9 0
1.6 0 0 0 1.1 0 0 0 0
1.6 0 0 0 1.1 0 0 1.0 0
17.5
0.4
0.5
1.1
0.5
2.3
18.2 23.1 2.7+
19.5 15.4 9.5
3.9 1.9 0
3.9 3.8 4.1
46.2 + 13.0 12.8
0 1.9 8.0
13.7
14.7
2.0
3.9
12.7
3.4
a
4.8 0 0 0 1.1 0 2.9 1.0 0
CyOv
a
@generation, # number of females, *significantly different from other SAM strains by chi-square test, +significantly different from other SAMP or SAMR strains by chi-square test, **a: p < 0.001; b: p < 0.01, Br: bronchopneumonia; In: interstitial pneumonia; Ab: abscess; Col: colitis; Amy: amyloidosis; CK: contracted kidney; Deg Tub: degeneratio of renal tubule; Inv: invagination; AN: angionecrosis; Ang: angiitis; TLB: thymic lymphoblastic lymphoma; NTLB: nonthymic lymphoblastic lymphoma; IB: immunoblastic lymphoma; FCC: follicular center cell lymphoma; Unc: unclassifiable lymphoma; HS: histiocyfic sarcoma; GL: granulocytic leukemia; Malig: malignant neoplasms; Cy Ov: ovarian cyst; Ben: benign neoplasms other than ovarian cyst.
120
Y. TAKEDA et a l
firmed in Congo red stained sections. Green birefringence of the Congo red strain under the polarizing microscope was considered to present amyloid (Takeshita et al., 1982). AA amyloid protein (secondary amyloidosis) was often observed in SAM mice with inflammatory lesions, particularly in SAMP2 and SAMP6. Senile amyloidosis, with apoA-1I protein deposits in organs or tissues, was observed more frequently and severely in SAMP1, -P2, -P7, -P9, -P10, and -P11 than in other strains of SAM. Contracted kidneys had a granular surface and were reduced in size according to the severity of the lesions. Destroyed nephrons were replaced by a proliferation of fibrous tissue. Degenerated renal tubules, "Deg Tub" in the table, showed vacuolar degeneration in their proximal convoluted sections. The vacuoles in the tubules did not show positive fat staining. Angionecrotic lesions were observed in the alimentary tract, pancreas, uterine adnexae, testes, ovaries, and kidneys. Angiitis was observed in the tongue, alimentary tract, pancreas, kidneys, uterine adnexae, and testes. Lymphoid cell neoplasms, histiocytic sarcoma, and granulocytic leukemia were diagnosed from gross and histological findings of the spleen, thymus, lymph nodes, bone marrow, and other affected organs, according to the classification of Pattengale and Taylor (1981, 1983). Three types of pathology in lymphoid cell neoplasms were observed in the SAM model: lymphoblastic lymphoma, immunoblastic lymphoma, and follicular center cell lymphoma. In this study, lymphoblastic lymphoma was classified into two types: thymic lymphoblastic lymphoma with definite neoplasm of thymic tissue in the anterior mediastinum, and nonthymic lymphoblastic lymphoma without any definite thymic neoplasm. Two lymphoid cell-derived neoplasms observed in a SAMP8 and a SAMR4 mouse are listed as " U n c " (unclassifiable) because of inadequate fixation (Table 1). Among the findings of routine postmortem examinations, amyloidosis, contracted kidney, immunoblastic lymphoma, histiocytic sarcoma, and ovarian cyst are age-associated pathologies the incidence of which increases with advancing age. The incidence of each pathologic finding was compared between SAMP and SAMR, among SAM strains, and also among SAMP or SAMR strains. The chi-square test on a two-by-two contingency table was used to test the statistical significance of differences between each genotype and all animals in the study as a ratio of animals with a pathology to those without incidence of certain common pathologic findings. As shown in Table 1, the incidence of amyloidosis (p < 0.001), contracted kidney (p < 0.001), and thymic lymphoblastic lymphoma (p < 0.001) was significantly higher in SAMP than in SAMR. On the other hand, colitis (p < 0.001), inflammatory changes other than pneumonia, abscess and colitis (p < 0.01), immunoblastic lymphoma (p < 0.001), histiocytic sarcoma (p < 0.001) and ovarian cyst (p < 0.001) were more frequent in SAMR than in SAMP. The incidence of bronchopneumonia and abscess did nol differ significantly between SAMP and SAMR. When the incidence of each abnormality was compared among all the SAM strains, that of amyloidosis and contracted kidney was significantly greater in SAMP2 and SAMP10, and was significantly lower in SAMR1, SAMR4, and SAMR5, than in the other genotypes of SAM. SAMP7 and SAMP9 had a significantly higher incidence of lymphoid cell neoplasms than did the others. A significantly lower incidence of lymphoid cell neoplasms was observed in SAMP10 than in the other strains of SAM. In the statistical analysis by the chi-square test of all the SAM strains, a p-value less than 0.001 was considered to be statistically significant. Among SAMP strains, SAMP7 (p < 0.001) and SAMP9 (p < 0.001) had a higher incidence of thymic lymphoblastic lymphoma than did the other genotypes. Among the SAMR strains, SAMR5 had a significantly lower incidence of immunoblastic lymphoma (p < 0.01) and lymphoid cell neoplasms (p < 0.01) than did the others. The incidence of ovarian cysts was significantly greater (p < 0.01) in SAMR1 than in the others. Table 2 shows a summary of the three most frequently seen autopsy findings in each SAM
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PATHOBIOLOGYFEATURESOF THE SAM MODEL
strain. In all the SAMP strains, amyloidosis was the most common autopsy finding with an incidence of 39.1-93.1% and was highest in SAMP10. The incidence in SAMR was 30.733.8%. Five of the SAMP strains, SAMP1, -P2, -P6, -P10, and -P11, had contracted kidney as the second most frequent autopsy finding; and in three, SAMP7, -P8, and -P9, it was lymphoid cell neoplasm. The third most common autopsy finding in seven strains of SAMP was abscess. In SAMP10 it was Deg Tub, degeneration of the renal tubules. Ovarian cyst (46.2%) and colitis (33.3%) were the most common pathologies in SAMR1 and SAMR5, respectively. The pathologic changes causing death were assessed in 1337 SAMP and 453 SAMR mice, in some by gross observation only. These included pneumonia, abscess, colitis, amyloidosis, contracted kidney, invagination, neoplasms, including lymphoid cell neoplasms, histiocytic sarcoma and other neoplasms, miscellaneous pathologic findings other than the above, and unknown causes. In the statistical analysis of the causes of death, the chi-square test was used. As shown in Table 3, colitis (p < 0.001), histiocytic sarcoma (p < 0.001), and unknown causes (p < 0.01) were more frequent causes of death in SAMR than in SAMP, and contracted kidney (p < 0.001) and lymphoid cell neoplasms (p < 0.001) were significantly more frequent in SAMP than in SAMR. In a comparison of the causes of death among all the SAM strains at the significance level of p < 0.001, pneumonia was a more frequent cause of death in SAMP3 and SAMR1 than in the other strains. In SAMP1 and SAMP10, contracted kidney was a more frequent cause of death than in the other genotypes. SAMP7, -P8, and -P9 mice died of lymphoid cell neoplasms significantly more frequently than did other strains of SAM. SAMR5 mice died more frequently without overt pathologic changes than did other strains of SAM. On the other hand, abscess in SAMP10, contracted kidney in SAMP7, -P8, -R1, and -R4, lymphoid cell neoplasms in SAMP2, -P6, -P10, and -P11, and unknown causes in SAMP7 and -P10 were less frequent causes of death than in the other strains of SAM. TABLE 2. TOP THREE AUTOPSYFINDINGS IN SAM STRAINS Strains
SAMP SAMP 1 SAMP2 SAMP3 SAMP6 SAMP7 SAMP8 SAMP9
1
Amy Amy Amy Amy
2
3
CK CK Br CK LN LN LN CK CK
Ab Ab Ab Ab Ab Ab Ab
Amy, Cy Ov
Ab
Ab Co
Amy Amy
HS Br Ab
Amy
Amy Amy
SAMP10
Amy
SAMPI 1 SAMR SAMR1 SAMR4 SAMR5
Amy
Deg Tub Br
Amy: amyloidosis; CK: contracted kidney; Ab: abscess; Br: bronchopneumonia; LN: lymphoid cell neoplasms; Deg Tub: degeneration of renal tubule; HS: histiocytic sarcoma; Cy Ov: ovarian cyst; Co: colitis.
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TABLE 3. AUTOPSY FINDINGSCAUSING DEATH(%)
Strains
Gen+
No.
Inflammation
Amy
CK
lnv
Neoplasms
Others
Unk
of mice
SAMPI SAMP2 SAMP3 SAMP6 SAMP7 SAMP8 SAMP9 SAMPI0 SAMPI1
SAMR
Ab
112 200 97 195 247 132 47 204 103
11.6 24.0 34.0* 15.9 22.7 15.2 6.4 1(I.8 24.3
22.3 23.5 20.6 20.0 16.6 18.2 19.2 6.8 13.6
0.9 3.5 0 2.1 0 4,5 0 1.0 0
0.9 0 0 2.6 0 0 0 0.5 2.9
28.6 21.5 3.1 17.4 7.7* (/.8" 8.5 47.5* 22.3
1337
17.2
t7.4
1.5
0.7
19.1
237 139 77
29.1" 23.0 16.9
17.7 15.8 10.4
2.5 14.4 1.3
0.8 0 1.3
453
25.2
15.9
6.0
0.7
67-8l 70-86 64-82 66-80 65-81 69-81 61-70 66-73 64-78
SAMP SAMRI SAMR4 SAMR5
Pn
59-73 60-71 55-70
p-value: SAMP vs. SAMR**
a
Col
a
LN
HS
ON
1.8 3.0 0 0 0 6.0 0 1,0 0
2.7 1.5" 6.2 0* 38.5* 23.5* 46.8* 1.5" 0*
0.9 0 0 0 0.4 0 0 0 0
1.8 0 0 0 0.8 0 0 0.5 0
11.6 9.5 12.4 14.3 3.6 8.3 8.5 20.1 8.7
17.0 23.5 23.7 27.7 9.7* 23.5 10.6 10.3" 28.2
1.3
12.2
0.1
0.4
10.9
18.9
3.0* 2.2* 1.3
t.3 0.7 0
5.5 10. t 2.6
4.6 6.5 5.2
3.0 2,1 2.6
9.7 5.0 16,9
22.8 20.1 41.5"
2.4
0.9
6.4
5.3
2,6
9,5
a
b
a
28.1
+generation; *significantly different from other SAM strains by ehi-square test; **a: p < 0.001, b: p < 0.01; Pn: pneumonia; Ab: abscess; Col: colitis; Amy: amyloidosis; CK: contracted kidney; lnv: invagination; LN: lymphoid cell neoplasms; HS: histocytic sarcoma; ON: other neoplasms; Unk: unknown.
The top three causes of death in each SAM strain are summarized in Table 4. Lymphoid cell neoplasm was the most common cause of death in SAMP7, -P8, and -P9; while contracted kidney was the most frequent cause of death in SAMP1 (28.6%) and SAMP10 (47.5%). The cause of death was unknown in 41.5% of SAMR5, in 28.2% of SAMP11, and in 27.7% of SAMP6. In these three, no known causes of death were of higher incidence. SAMR5, a strain with normal aging, had fewer fatal pathologies such as lymphoid cell neoplasms and histiocytic sarcoma, contracted kidney, or inflammatory changes (except for colitis) than did the other strains (Table 1). Interestingly enough, amyloidosis with the highest or second highest incidence in autopsy findings in all SAM strains, was only a minor cause of death (Tables 1-4), because other more severe pathologies that often coexisted with amyloidosis, such as malignant neoplasms, contracted kidney, etc., were the causes of death. PATHOBIOLOGICAL FEATURES REVEALED BY SYSTEMATICALLY DESIGNED STUDIES Besides the routine postmortem examinations, several studies were conducted to assess systematically and dynamically various age-related pathologic changes. A series of behavioral studies on SAMP8 and SAMP10 disclosed age-related deficits in learning and memory (Miyamoto et al., 1986; Yagi et al., 1988; Ohta et al., 1989; Flood and Morley, 1992, 1993; Shimada et al., 1993). In SAMP10, age-related spontaneous brain atrophy was also observed. Miyamoto et al. (1992) reported a characteristic emotional disorder, reduced anxiety-like behavior, and a profound disorder of circadian rhythm, in addition to the deficits in learning and memory, in SAMP8/Ta reared under specific pathogen-free (SPF) conditions at Takeda Chemical Industries, Ltd. To clarify the pathogenic mechanisms in the learning and memory deficits
PATHOBIOLOGY FEATURES OF THE SAM MODEL
123
TABLE4. TOPTHREECAUSESOFDEATHIN SAM STRAINS Strains
SAMP SAMP1 SAMP2 SAMP3 SAMP6 SAMP7 SAMP8 SAMP9 SAMP10 SAMP11 SAMR SAMRI SAMR4 SAMR5
1
2
3
CK Pn Pn Unk LN LN LN CK Unk
Ab Ab Unk Ab Pn Unk Ab Others Pn
Unk Unk Ab CK Ab Ab Unk Pn CK
Pn Pn Unk
Unk Unk Pn
Ab Ab Others
CK: contractedkidney;Ab: abscess;Unk:unknown;Pn: pneumonia, including bronchopneumoniaand interstitial pneumonia; LN: lymphoidcell neoplasms; Others: see text. in SAMP8 and SAMP10, a series of neuropathological, neurochemical, and neuropharmacological experiments were performed (Shimada et al., 1992; Nomura et al., 1994; Flood and Morley, 1994). In the course of development of SAM, we became aware of spontaneous leg fractures in a few aged SAM mice. Therefore, we began to examine systematically the age-related changes in bone in several strains of SAM and found that SAMP6 mice showed senile osteoporosis characterized by a low peak bone mass (Matsushita et al., 1986). A series of studies to clarify the pathogenesis of osteoporosis in SAMP6 is now under way in our laboratory (Tsuboyama et al., 1989a,b, 1993; Takahashi et al., 1994; Okamoto et al., 1995) as well as in others (Kawase et al., 1989; Suda et al., 1994). A systematic study on the condyle of the temporomandibular joint in several strains of SAM revealed that all strains of SAM develop degenerative joint disease. However, the short-lived SAMP strains develop degenerative changes earlier than do the SAM strains. SAMP3 mice are the first to show degenerative joint changes (approximately 50% at seven to nine months of age and 100% over 12 months of age) and thereafter have the highest incidence of severe changes with overt deformity (Chen et al., 1989). Examinations of the eyes and ears showed age-related cataracts in SAMP9 and age-related hearing impairment in SAMP1 and SAMR1. Persistence of the hyaloid vascular system is a necessary, but not the sole, factor in SAMP9 cataracts (Hosokawa et al., 1988, 1993). The persistence of the hyaloid vascular system might affect the breakdown of the lens capsule, a trigger of cataract formation, and subsequently lead to increased Ca content and transglutaminase activity in the lens to reduce the solubility of [~-crystallins (Ashida et al., 1994). SAMP1 and SAMR1 showed an age-related auditory loss expressed as elevated thresholds, prolongation of interpeak intervals I-III and 1-IV, and decreased amplitude of wave I assessed by auditory brainstem responses (ABRs) (Saitoh et al., 1994). SAMP1 showed a more rapid and
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severe auditory loss with age than did SAMRI. Morphological studies showed an age-related decrease in cell density as well as the size of spiral ganglion neurons in SAMPI and SAMR1 (Saitoh et al., 1994). An age-related loss of the inner and outer hair cells was observed in both SAMP1 and SAMRI. The changes in the spiral ganglion neurons and hair cells, however, appeared earlier and progressed more rapidly in SAMP1 than in SAMR1. Furthermore, SAMPI showed greater age-related atrophy of the stria vascularis than did SAMR1. These results suggest that hearing impairment in SAM is due to a combination of sensory and strial presbycusis as well as to neural presbycusis (Saitoh et al., 1995). Morphometric and physiological studies of the respiratory system showed senile hyperinflation of the lungs in SAMP1 and SAMR 1. Histologic observation, however, revealed no evidence of destruction of the alveolar wall or elastic fibers in the lungs. The hyperinflationary changes occurred similarly in both SAMPI and SAMR1, but in an accelerated manner in SAMPI ; and, finally, senile hyperinflation of the lungs occurred in the senescence phase in both strains (Kurozumi et al., 1994). A series of immunobiological studies was performed. In vivo experiments showed that primary antibody response and delayed-type hypersensitivity (DTH) reaction to sheep red blood cells (SRBC) were significantly more impaired by six months of age in SAMP1 and SAMP2 than in SAMR1 and ordinary strains such as C3H/He and B10.BR (Mitsuoka and Hanada, 1988). To clarify the nature of the immunobiological defects observed in vivo, in vitro experiments were conducted, and it became evident that SAMP1 mice had a profound defect in antibody response to a T-dependent antigen, SRBC as early as two months of age and a negligible response at a later age (Hosokawa et al., 1987a). This impaired antibody response was considered to be closely related to dysfunction of T-helper cells, while other T-cell activities such as mixed lymphocyte reaction, cytotoxic T lymphocyte generation and DTH reaction remained normal. B-cell and antigen-presenting cell activities were also normal (Hosokawa et al., 1987b). Recently, Abe e t al. (1994) reported that a helper T-cell defect similar to that in SAMP1 was found in SAMP8 mice that show characteristic deficits in learning and memory with advancing age. Furthermore, low natural killer (NK) cell activity was observed in SAMP8. Among the pathobiological findings in SAM revealed by routine postmortem examinations and also by systematic studies, there are pathologic phenotypes that are often characteristic enough to distinguish among all the SAM strains, as shown in Table 5. Furthermore, most of the "age-dependent" geriatric disorders seen in humans, which were described as "inevitable disorders as a direct consequence of physiological senescence" by Cotran e t al. (1989), are included in the pathobiological phenotypes in SAM: these are osteoporosis, degenerative joint disease, cataract, hyperinflation of lungs, and hearing impairment. The evidence provides support for our proposal that the SAM model is a valid one for research on aging. CONCLUSIONS In general, one of the most important issues in the development and maintenance of animal models for biomedical research is the maintenance of the expression of the characteristic phenotypes. The SAM model, the characteristics of which have been strictly monitored pathobiologically as well as genetically (Festing, 1992, 1993), is expected to clarify the mechanisms of age-associated pathologies and to provide effective methods of modulating or ameliorating the advance of senescence and the disease processes involved in age-associated pathologies. The fundamental mechanisms of accelerated, as well as primary, aging processes can be demonstrated in the SAM model.
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TABLE 5. PATHOBIOLOGICALPHENOTYPESIN SAM MICE Strains SAMP SAMP1 SAMP2 SAMP3 SAMP6 SAMP7 SAMP8 SAMP9 SAMP10 SAMP11 SAMR SAMR1 SAMR4 SAMR5
Phenotypes
Senile amyloidosis, contracted kidney, impaired immune response, hyperinflation of lungs, hearing impairment. Senile and secondary amyloidosis, contracted kidney, impaired immune response. Degenerative temporomandibular joint disease. Senile osteoporosis, secondary amyloidosis. Thymic lymphoblastic lymphoma, senile amyloidosis. Deficits in learning and memory, emotional disorder (reduced anxiety-like behavior),* impaired immune response. Cataract, thymic lymphoblastic lymphoma, senile amyloidosis. Brain atrophy, deficits in learning and memory, emotional disorder (depressive behavior),* contracted kidney, senile amyloidosis. Contracted kidney, senile amyloidosis. Nonthymic lymphoma,t histiocytic sarcoma, ovarian cyst. Nonthymic lymphoma,t histiocytic sarcoma. Colitis
*Reported in SAMP8/'ra or SAMP10/Ta mice reared under SPF conditions at Takeda Chemical Industries Ltd. tlncludes nonthymic lymphoblastic lymphoma, immunoblastic lymphoma and follicular center cell lymphoma. Acknowledgments--This work was supported in part by grants from the Ministry of Education, Science and Culture and the Ministry of Health and Welfare, Japan. Gratitude is extended to Drs. W. H. Chen, A. Ohta, and N. Seriu, and K. Kadota and K. Yasuoka for technical assistance. We are also grateful to Dr. A. Cary for critical reading of the manuscript. REFERENCES ABE, Y., YUASA, M., KAJIWARA, Y., and HOSONO, M. Deficits of immune cells in the Senescence-Accelerated Mouse: A model for learning and memory deficits in the aged. Cell Immunol. 157, 59-69, 1994. ASHIDA, Y., TAKEDA, T., and HOSOKAWA, M. Protein alterations in age-related cataract associated with a persistent hyaloid vascular system in senescence-accelerated mouse (SAM). Exp. Eye Res. 59, 467-474, 1994. CHEN, W.H., HOSOKAWA, M., TSUBOYAMA, T., ONO, T., IIZUKA, T., and TAKEDA, T. Age-related changes in the temporomandibular joint of the Senescence Accelerated Mouse (SAM): SAM-P/3 as a new murine model of degenerative joint disease. Am. J. Pathol. 135, 379-385, 1989. COTRAN, R.S., KUMAR, Y., and ROBBINS, S.L. Disease of aging. In: Robbins Pathologic Basis of Disease, pp. 543-551, W.B. Saunders Company, Philadelphia, PA, 1989. FESTING, M.F.W. Origins and characteristics of inbred strains of mice. Mouse Genome 90, 236-237, 1992. FESTING, M.F.W. Origins and characteristics of inbred strains of mice. Mouse Genorne 91, 471-472, 1993. FLOOD, J.F. and MORLEY, J.E. Early onset of age-related impairment of aversive and appetitive learning in the SAM-P/8 mouse. J. Gerontol. 47, B52-B59, 1992. FLOOD, J.F. and MORLEY, J.E. Age-related changes in foot shock avoidance acquisition and retention in senescence accelerated mouse (SAM). Neurobiol. Aging 14, 153-157, 1993. FLOOD, J.F., and MORLEY, J.E. Age-related changes in the pharmacological improvement of retention in SAM8 mice. In: The SAM Model of Senescence, Takeda, T. (Editor), pp. 89-94, Excerpta Medica, Elsevier Science B.V., Amsterdam, 1994. HIGUCHI, K., MATSUMURA, A., HONMA, A., TAKESHITA, S., HASHIMOTO, K., HOSOKAWA, M., YASUHIRA, K., and TAKEDA, T. Systemic senile amyloid in Senescence-Accelerated Mice: A unique fibril protein demonstrated in tissues from various organs by the unlabeled immunoperoxidase method. Lab. Invest. 48, 231-240, 1983. HIGUCHI, K., NAIKI, H., KITAGAWA, K., HOSOKAWA, M., and TAKEDA, T. Mouse senile amyloidosis: ASsAM
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changes in SAMP6 are due to a decrease in osteoblast progenitor cells. In: The SAM Model of Senescence, Takeda, T. (Editor), pp. 47-52, Excerpta Medica, Elsevier Press B.V., Amsterdam, 1994. TAKAHASHI, K., TSUBOYAMA, T., MATSUSHITA, M., KASAI,R., OKUMURA, H., YAMAMURO, T~, OKAMOTO, Y., TORIYAMA, K., KITAGAWA, K., and TAKEDA, T. Modification of strain-specific femoral bone density by bone marrow derived factors administered neonatally: A study on the spontaneously osteoporotic mouse, SAMP6. Bone Miner. 24, 245-255, 1994. TAKEDA, T., HOSOKAWA, M., TAKESHITA, S., IRINO, M., HIGUCHI, K., MATSUSHITA, T., TOMITA, Y., YASUHIRA, K., HAMAMOTO, H., SHIMIZU, K., ISHII, M., and YAMAMURO, T. A new murine model of accelerated senescence. Mech. Ageing Dev. 17, 183-194, 1981. TAKEDA, T., HOSOKAWA, M., and HIGUCHI, K. Senescence-Accelerated Mouse (SAM): A novel murine model of accelerated senescence. J. Am. Geriatr. Soc. 39, 911-919, 1991. TAKEDA, T., HOSOKAWA, M., HIGUCHI, K., HOSONO, M., AKIGUCHI, I., and TAKEDA, T. A novel murine model of aging, Senescence-Accelerated Mouse (SAM). Arch. Gerontol. Geriatr. 19, 185-192, 1994. TAKESHITA, S., HOSOKAWA, M., IRINO, M., HIGUCHI, K., SHIMIZU, K., YASUHIRA, K., and TAKEDA, T. Spontaneous age-associated amyloidosis in Senescence-Accelerated Mouse (SAM). Mech. Ageing Dev. 20, 13-23, 1982. TSUBOYAMA, T., TAKAHASHI, K., MATSUSHITA, M., OKUMURA, H., YAMAMURO, T., UMEZAWA, M., and TAKEDA, T. Decreased endosteal formation during cortical bone modelling in SAM-P/6 mice with a low peak bone mass. Bone Miner. 7, 1-12, 1989a. TSUBOYAMA, T., MATSUSHITA, M., OKUMURA, H., YAMAMURO, T., HANADA, K., and TAKEDA, T. Modification of strain-specific femoral bone density by bone marrow chimerism in mice: A study on spontaneously osteoporotic mouse (SAM-P/6). Bone 10, 269-277, 1989b. TSUBOYAMA, T., TAKAHASHI, K., YAMAMURO, T., HOSOKAWA, M., and TAKEDA, T. Cross mating study on bone mass in the spontaneously osteoporotic mouse (SAM-P/6). Bone Miner. 23, 57--64, 1993. YAGI, H., KATOH, S., AKIGUCHI, I., and TAKEDA, T. Age-related deterioration of ability of acquisition in memory and learning in Senescence Accelerated Mouse: SAM-P/8 as an animal model of disturbances in recent memory. Brain Res. 474, 86--93, 1988.