To address specifically how the SARS-CoV-2
PAMPs affect SnCs, senescent human kidney
endothelial cells (fig. S4) were treated with
pyrogen-free recombinant S1 protein. Exposing
endothelial SnCs to S1 for 24 hours signifi-
cantly increased secretion of the majority of
endothelial SASP factors measured in the con-
ditioned media (composite scoreP< 0.0089
comparing SnCs to non-SnCs) (Fig. 2A and
table S2). Similar albeit less distinctive results
were obtained by using kidney endothelial cells
in which senescence was induced through rep-lication rather than radiation (fig. S5 and table
S3). In addition, treatment of human sub-
cutaneous adipocyte progenitor SnCs with S1
increased expression of the key preadipocyte
SASP factors,IL1aandIL1b, at the mRNA
level (fig. S6). Consistent with the LPS data,Camellet al.,Science 373 , eabe4832 (2021) 16 July 2021 4 of 12
Fig. 3. Old mice are vulnerable
to a NME that includes acute
mouseb-coronavirus infection.
(A) Young (3-month-old) and old
(20- to 24-month-old) WT mice
were exposed to NME bedding
produced from pet store mice for
7 days. Survival was monitored for
35 days after initiation of NME
(n= 10 young;n= 18 old). Log-
rank (Mantel Cox) test. (B) Gene
expression in three tissues of SPF
or NME (6- to 7-day exposure)
young and old mice (n= 3 young
SPF;n= 5 old SPF;n= 14 young
NME;n= 13 old NME) measured
with quantitative PCR. Expression
was normalized to young SPF
mice. Means ± SEM, two-way
ANOVA and post hoc comparison
Tukey’s honestly significant
difference were used to compare
the two animal cohorts within a
treatment group. P< 0.05, P<
0.01, P< 0.001, **P<
0.0001. Arrows and asterisks:
gray, SPF old versus young; black,
NME old versus young; red, old
SPF versus old NME. (C) Serum
cytokine levels in young and
old mice (n=3youngSPF;
n=5oldSPF;n= 19 young NME;
n= 17 old NME) measured with
ELISA at day 5 after NME. Statistics
are as described in (B). (D) Serol-
ogy to detect antibodies against
microbes in NME bedding. (Right)
The mouse pathogens commonly
tested for by Charles River Labora-
tory to define SPF housing. The pie
charts illustrate the exposures
detected in individual young and old
mice (n= 24 young;n=21to
23 old) day 11 after initiation of
NME. Serology of pet store mice is
illustrated below. (E) Representative
images of hematoxylin and eosin
(H&E) staining or MHV immuno-
histochemistry in liver sections from
young and old mice exposed to
NME. (F) (Top) Schematic to illus-
trate the experimental design.
Young (6-month-old) or old (22-month-old) female mice were inoculated with a sublethal dose of MHV. Thirty days later, naïve and inoculated mice were exposed to NME
bedding for 3 weeks. (Bottom) Serum antibodies against three different MHV antigens measured 21 days after MHV inoculation and reported as relative scores. The
dotted line indicates the limit of detection (LOD). Means ± SEM, unpaired two-tailed Student’sttest. P< 0.01, ****P< 0.0001. (G) Survival of MHV-inoculated and
naïve mice measured for 42 days after initiation of NME. Log-rank (Mantel Cox) test.
0 10 20 30 40020406080100Days (after NME initiation)% SurvivalMHV-Young
(n=26)MHV-Old
(n=24)Naive Old
(n=10)Antibody (day 11)
Young OldPetstoreMHV
MVTM
MPV-1
MPV-2
NS-1
EDIM
MNV
GDVII
MPUL
NoneYoungOldH&E MHV IHCRel. expressionLungRel. expressionLiver KidneyABDEFGp16Ink4ap21Cip130MHV-A59
6m22mMHVA590 10 20 30 400255075100DaysSurvival(%)1p<0.0001Young
OldDay 0 51 70015
10
520NMENMENMEp<0.0001024602468040608020
0124
3502030405010SPF NME SPF NME SPF NMESPF NME SPF NME SPF NMESPF NME SPF NME SPF NMESPF NME SPF NME SPF NMESPF NME SPF NME SPF NMERel. expressionIl6Rel. expressionMcp1Rel. expression
Tnf01020304002468015
10
520040608020
040608020
015
10
520015
10
52002460203040501002030405010
01000200030000510400 15
300
200
100
0
SPF NME SPF NME SPF NME SPF NMEpg/mLCYoung
OldYoungOldIL-6 IL-10 CCL11 TNFα**** ******0 LOD201030MHV Antibody Scores*
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