Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2021 Nov 25;4(1):1326.
doi: 10.1038/s42003-021-02843-2.

Osteoblastic Swedish mutant APP expedites brain deficits by inducing endoplasmic reticulum stress-driven senescence

Jin-Xiu Pan #  1   2 , Dong Sun #  1 , Daehoon Lee  1   2 , Lei Xiong  1   2 , Xiao Ren  1 , Hao-Han Guo  1 , Ling-Ling Yao  1 , Yuyi Lu  1 , Caroline Jung  1 , Wen-Cheng Xiong  3   4
Affiliations

Osteoblastic Swedish mutant APP expedites brain deficits by inducing endoplasmic reticulum stress-driven senescence

Jin-Xiu Pan et al. Commun Biol. .

Abstract

Patients with Alzheimer's disease (AD) often have osteoporosis or osteopenia. However, their direct link and relationship remain largely unclear. Previous studies have detected osteoporotic deficits in young adult Tg2576 and TgAPPsweOCN mice, which express APPswe (Swedish mutant) ubiquitously and selectively in osteoblast (OB)-lineage cells. This raises the question, whether osteoblastic APPswe contributes to AD development. Here, we provide evidence that TgAPPsweOCN mice also exhibit AD-relevant brain pathologies and behavior phenotypes. Some brain pathologies include age-dependent and regional-selective increases in glial activation and pro-inflammatory cytokines, which are accompanied by behavioral phenotypes such as anxiety, depression, and altered learning and memory. Further cellular studies suggest that APPswe, but not APPwt or APPlon (London mutant), in OB-lineage cells induces endoplasmic reticulum-stress driven senescence, driving systemic and cortex inflammation as well as behavioral changes in 6-month-old TgAPPsweOCN mice. These results therefore reveal an unrecognized function of osteoblastic APPswe to brain axis in AD development.

PubMed Disclaimer

Conflict of interest statement

The authors declare no competing interests.

Figures

Fig. 1
Fig. 1. Specific expression of APPswe in OB-lineage cells in TgAPPsweOCN mice.
a Illustration of the transgene and generation of the conditional transgenic mice selectively expressing human APPswe in an OCN-Cre dependent manner. b, c Western blot analysis of human APP (hAPP) protein levels in BMSCs, hippocampus, and cortex of 6-MO control (LSL-APPswe) and TgAPPsweOCN mice. b Representative blots; and c quantification. d, e ELISA analyses of human Aβ40(d) and Aβ42(e) levels in serum, BMSCs (50 μg in total protein), and brain homogenates including hippocampus and cortex (300 μg total protein) from 6-MO control, TgAPPsweOCN, and Tg2576 mice. f RT-PCR analysis of hAPP gene expression in BMSCs, olfactory bulb, cerebellum, hippocampus, and cortex of 6-MO control and TgAPPsweOCN mice. g RT-PCR analysis of Cre expression in BMSCs, hippocampus, and cortex of 6-MO control (LSL-APPswe) and TgAPPsweOCN mice. All data were presented as mean ± SD. *p < 0.05, **p < 0.01, ***p < 0.001 (n = 3 mice). Mann–Whitney U test was used in c and g, and one-way ANOVA followed by Tukey post hoc test was used in d–f.
Fig. 2
Fig. 2. Elevated reactive astrocytes, microglial cells, and inflammatory cytokines in 6-MO TgAPPsweOCN cortex, but not hippocampus.
a Representative images of co-immunostaining with IBA1 (green), GFAP (magenta), and DAPI (blue) of hippocampal sections from 6-MO control (LSL-APPswe) and TgAPPsweOCN mice. Scale bars: 200 µm (upper) and 20 µm (lower). b Quantification of data in a. c Representative images of co-immunostaining with IBA1 (green), GFAP (magenta), and DAPI (blue) of cortex sections from 6-MO control (LSL-APPswe) and TgAPPsweOCN mice. Scale bars: 100 µm (upper) and 20 µm (lower). d Quantification of data in c. e Representative Western blots using antibodies against hAPP, GFAP, and IBA1 in homogenates of cortex and hippocampus of control and TgAPPsweOCN mice. GAPDH was used as a loading control. f Quantification of the data in e. g–h Real-time PCR (RT-PCR) analysis of indicated gene expressions in 6-MO control (LSL-APPswe) and TgAPPsweOCN cortex (g) and hippocampus (h). All quantification data were presented as mean ± SD (n = 3–4). *p < 0.05, **p < 0.01, ***p < 0.001. Student’s t test was used in b, d, and f–h.
Fig. 3
Fig. 3. Age-dependent anxiety- and depression-like behaviors in TgAPPsweOCN mice.
a, b OFT: Representative tracing images (a), and quantifications of total distance and center duration time (b) were shown. c, d EPMT: Representative tracing images (c), and quantifications of open arm duration time and entries (d) were shown. e LDT: Quantifications of the time spent in the light room and the number of transitions into the light room. f TST, g FST, and h SPT. In all these behavior tests, 6-MO and 12-MO control (LSL-APPswe) and TgAPPsweOCN mice (males) were examined. All quantification data were shown as mean ± SD (n = 10 mice). *p < 0.05, **p < 0.01, ***p < 0.001, Student’s t test.
Fig. 4
Fig. 4. Age-dependent alterations in spatial learning and memory in TgAPPsweOCN mice.
a–c 3-MO control (LSL-APPswe) and TgAPPsweOCN male mice were subject to Morris water maze (MWM) (a, b) and Novel Object Recognition (NOR) (c) tests. d–f 6-MO control (LSL-APPswe) and TgAPPsweOCN male mice were subject to MWM (d, e) and NOR (f) tests. g–i 12-MO control (LSL-APPswe) and TgAPPsweOCN male mice were subject to MWM (g, h) and NOR (i) tests. In MWM tests, the latencies to reach the hidden platform during the training period were showed in a, d, and g; and the representative tracing images and quantification of time spent in target quadrant, platform crossing time, and swim speed were shown in b, e, and h. In NOR tests, the time spent with novel object per total time with both objects as the novel object preference was quantified, shown in c, f, and i. All values were presented as mean ± SD (n = 10 mice). *p < 0.05, one-way ANOVA followed by Tukey post hoc test was used in a, d, and g, and Student’s t test was used in b, c, e, f, h, and i.
Fig. 5
Fig. 5. Increased cytokines and chemokines in APPswe+ OB-progenitor cells.
a Schematic of purification and RNA-seq of Tdtomato+ (Td+) OB progenitors from control (OCN-Cre; Ai9) and TgAPPsweOCN; Ai9 mice. b–d Volcano plots (b), GO analysis of up/down-regulated genes (c), and heat map (d) of differentially expressed genes identified by RNA-seq. e RT-PCR analysis of AD risk gene App, Vps35, Trem2, Apoe, Ptk2b, and Sorl1; bone-mass regulator Sp7, Nfatc1, Col1a1, Spp1, Sparc, Bmp2, Lrp4, and Ctnnb1; cytokine Il1b, Il6, and Il10; chemokine Ccl5 and Cxcl1, growth factor Tgfb1 gene expression in purified Td+ OB progenitors from 6-MO control (OCN-Cre; Ai9) and TgAPPsweOCN; Ai9 mice. All values were presented as mean ± SD (n = 3 mice). *p < 0.05, **p < 0.01, and ***p < 0.001, by Mann–Whitney U test.
Fig. 6
Fig. 6. Increased cellular senescence in APPswe+ OB-lineage cells.
a SA-β-gal staining of 3-MO and 6-MO BMSCs from control (LSL-APPswe) and TgAPPsweOCN mice. Scale bar, 20 µm. b Quantification of SA-β-gal+ cell densities (mean ± SD; n = 3 independent experiments). **p < 0.01, ***p < 0.001. c Western blot analysis of indicated protein expression in BMSCs from mice with indicated genotypes (at 6-MO). GAPDH was used as a loading control. d Quantification analyses of the data in c, *p < 0.05, ***p < 0.001. mean ± SD n = 3. Mann–Whitney U test.
Fig. 7
Fig. 7. Diminished behavior phenotypes in TgAPPsweOCN mice treated with senescence inhibitors.
a Schematic diagram of experimental design. 6-MO control (LSL-APPswe, n = 10 males) and TgAPPsweOCN mice were treated with Veh (10%PEG 400) (n = 10 males) or DQ (D 5 mg/kg, Q 50 mg/kg, dissolved in 10% PEG 400, once per two weeks) (n = 9 males), starting at age of 3-MO, and then subjected to indicated behavior tests at 6-MO. b OFT: Representative tracing images and quantifications of the total distance and the center duration time were shown. n.s. not significant, *p < 0.05. c EPMT: Representative tracing images and quantifications of the open arm duration time and entries were shown. *p < 0.05, **p < 0.01, ***p < 0.001. d LDT: Quantifications of the time spent in the light room and the number of transitions into the light room were shown. n.s. not significant, *p < 0.05. e TST, f FST, and g SPT were shown. *p < 0.05, **p < 0.01. h–i MWM: the latency to reach the hidden platform during the training period (h), and representative tracing image and quantification of the time spent in the target quadrant, platform crossing time and swim speed (i) were shown. *p < 0.05, **p < 0.01. One-way ANOVA followed by Tukey post hoc test. All data were presented as mean ± SD.
Fig. 8
Fig. 8. Increased cytokines and chemokines in TgAPPsweOCN serum samples.
a Representative images of serum L-Series label-multiplex antibody arrays of ~7-MO control and TgAPPsweOCN mice. b Volcano plots analysis of a. c Heat map of data in a. n = 4, significant difference was set at p < 0.05. d Elisa assays of serum IL1β and IL6 levels in ~7-MO control and TgAPPsweOCN mice. The data were presented as mean ± SD (n = 4 mice). *p < 0.05 by Student’s t test. e Comparison between this antibody array with secreted factors by RNA-seq of purified Tdtomato+ BMSCs. f Comparison of the changes (upregulated secreted proteins in Tg2576 over control mice) to those detected in TgAPPsweOCN mice.
Fig. 9
Fig. 9. APPswe induction of OB-senescence via ER stress.
a Heat map of differentially expressed ER stress or anti-stress related genes identified by RNA-seq in control (OCN-Cre; Ai9) and TgAPPsweOCN; Ai9 Td+ OB-progenitors (detail analysis was described in Methods). b RT-PCR analysis of ER stress-related genes Grp78, Atf6, Hsp90b1, Eif2ak3, Ern1, Hsp90aa1, and Hspa2 and anti-stress related gene Sirt3 gene expression in purified Td+ BMSCs from 6-MO control (OCN-Cre; Ai9) and TgAPPsweOCN; Ai9 mice, *p < 0.05, **p < 0.01, ***p < 0.001, mean ± SD, n = 3, Mann–Whitney U test. c Western blot analysis of indicated protein expression in BMSCs from mice with indicated genotypes (at 6-MO). GAPDH was used as a loading control. d Quantification of data in c, *p < 0.05, **p < 0.01. mean ± SD, n = 4, Student’s t test. e Western blot analysis of indicated protein expression in BMSCs from 6-MO control and TgAPPsweOCN with or without 0.25 mM 4-PBA (4-Phenylbutyric acid) treatment. f Quantification analyses of the data in e, *p < 0.05, n = 3. g SA-β-gal staining of 6-MO control and TgAPPsweOCN BMSCs with vehicle (Veh)(PBS) and 4-PBA treatment, respectively, scale bar, 20 µm. h Quantification of SA-β-gal+ cell densities in g (mean ± SD; n = 5, **p < 0.01, ***p < 0.001). Two-way analysis of variance test was used in f and h.
Fig. 10
Fig. 10. Summary and working hypothesis for APPswe in OB-lineage cells to regulate brain-pathology and behavior changes.
a Summary of phenotypes detected in TgAPPsweOCN mice at indicated ages. b Illustration of the working model.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

References

    1. Leng, F. & Edison, P. Neuroinflammation and microglial activation in Alzheimer disease: where do we go from here? Nat. Rev. Neurol.17, 157–172 (2020). - PubMed
    1. Wang J, Gu BJ, Masters CL, Wang YJ. A systemic view of Alzheimer disease—insights from amyloid-beta metabolism beyond the brain. Nat. Rev. Neurol. 2017;13:612–623. - PubMed
    1. Basgoz B, Ince S, Safer U, Naharci MI, Tasci I. Low bone density and osteoporosis among older adults with Alzheimer’s disease, vascular dementia, and mixed dementia: a cross-sectional study with prospective enrollment. Turkish J. Phys. Med. Rehabil. 2020;66:193–200. - PMC - PubMed
    1. Mjoberg B, Hellquist E, Mallmin H, Lindh U. Aluminum, Alzheimer’s disease and bone fragility. Acta Orthopaedica Scandinavica. 1997;68:511–514. - PubMed
    1. Frame G, Bretland KA, Dengler-Crish CM. Mechanistic complexities of bone loss in Alzheimer’s disease: a review. Connect. Tissue Res. 2020;61:4–18. - PubMed

Publication types