Introduction

Alzheimer’s disease (AD) is the most common cause of dementia in the elderly, which affects over 50 million people worldwide [1, 2]. Intraneuronal formation of neurofibrillary tangles (NFTs) composed of hyperphosphorylated tau protein is a pathological hallmark of AD [3, 4] and is well recognized to correlate with cognitive deficits of patients [4, 5]. Therefore, increasing attention has been paid on the mechanisms underlying how tau pathology contributes to AD, as well as the tau-targeted drug discovery [6,7,8,9]. However, studies of tau pathology and drug development are largely limited by the lack of readily available mouse models that mimic the AD-specific tau pathology.

Several tau transgenic (Tg) mouse models have been used for decades in the research of tau pathology [10], including: (1) lines overexpressing wild-type (WT) or mutant human tau (hTau), such as ALZ17 (WT) [11] and PS19 (P301S) [12]; (2) Tg mice with expression of full-length WT or mutant hTau in the absence of endogenous murine tau, such as the hTau line (JAX005491) [13,14,15]; and (3) mice with expression of WT or mutant full-length hTau under the induction of tetracycline, such as rTg4510 (CaMK2a:P301L) [16] and rT2 (Col1a1:P301L) [17]. However, these Tg lines show limitations in imitating the tau pathology in AD [18]. First, most non-inducible hTau-Tg lines do not exhibit overt phosphorylated tau accumulation in the brain until 6–9 months of age [11, 19], so researchers have to spend much time and resources to keep mice before experiments. Second, the commonly studied tau mutations in tau lines such as P301L and P301S are only found in frontotemporal dementia but not in AD patients [20, 21]. In addition, some other mutated tau lines like rTg4510 (CaMK2a:P301L) show serious motor impairments, which may disturb the evaluation of cognitive functions in behavioral tests [17, 22,23,24].

To generate an easy-to-use animal model with accurate simulation of the actual AD tau pathology, we generated a novel hTau368-Tg mouse line, in which the AD-like truncated hTau N1–368 (hTau368) was expressed under the neuron-specific enolase 2 (Eno2) promoter with induction of tetracycline (tet-on). The neurotoxic tau fragment N1–368 is cleaved by asparagine endopeptidase (AEP), and it accumulates and mediates NFT formation during aging and AD [25]. Recently, we have used hTau368 mice as a model to test the effectiveness of a peptide drug designed to specifically facilitate tau dephosphorylation [26]. For better use of this model, we here report more detailed characterization of AD-like pathologies in this mouse line.

Materials and methods

Animals

All mice were housed in groups of three to four per cage, under a 12 h light/dark cycle at 23–25 °C. Food and water were available ad libitum. Doxycycline hyclate (Beyotime, ST0398) was dissolved in drinking water (2 mg/l) and administered ad libitum. Equal numbers of male and female mice were randomly assigned to vehicle (Veh) or dox-treatment (Dox) groups. Mice were sacrificed at age of 7 months unless otherwise specified. All experiments and data analyses were conducted by experimenters blind to the grou**s. All animal experiments were conducted in accordance with relevant ethical regulations for animal testing and research, and were approved by institutional guidelines and the Animal Care and Use Committee of Tongji Medical College, Huazhong University of Science and Technology.

Tg hTau368 generation

The hTau368 mice were generated jointly by our laboratory and Nan**g Biomedical Research Institute of Nan**g University. Specifically, an Eno2-hMAPT vector, Insulator-pTRE3G-Kozak-ATG-hMapt N1-368-CDS-TGA-polyA-bGH-polyA-TAA-rtTA3G-Kozak-ATG-Rat Eno2 promoter-Insulator, was constructed by linking reactive cloning. pTRE is a tetracycline-induced promoter. hMAPT is the targeted gene searched through https://www.ncbi.nlm.nih.gov/gene query (Gene ID: 4137). MAPT contains 12 transcripts, of which the 2N4R transcript NM_001123066.4 and the sequences encoding tau N1-368 were selected. PolyA was used as the termination signal. rtTA3G was the tetracycline-regulated transcription activator. Rat Eno2 promoter (Gene ID: 24,334, transcript xM_0062373304) was used to control the expression of rtTA3G.

The Eno2-hMAPT vector was confirmed by DNA sequencing and microinjected into the nucleus of fertilized eggs of C57BL/6 J mice. Surviving fertilized eggs were implanted into the uterus of pseudo-pregnant C57BL/6 J mice. Founder mice in the F0 offspring were identified by PCR at 1 month old. Whole-genome sequencing was performed to find the insertion site of the targeted gene (Additional file 1: Fig. S1). Identification of homozygous and hemizygous hTau368 was conducted by PCR using primers shown in Additional file 1: Table S1. F1 hTau368 mice were used for expand reproduction and F2–4 offspring with transgene were used for experiments in this study. C57BL/6 J mice were used as wild-type controls.

Antibodies

Antibodies used in the present study are summarized in Additional file 1: Table S2. The polyclonal rabbit anti-TauN368 antibody gifted by Professor Keqiang Ye was developed in his laboratory and had been reported previously [25]. The new monoclonal mouse anti-tauN368 antibody was jointly developed by AtaGenix (Wuhan, China). In brief, a keyhole limpet hemocyanin (KLH)-conjugated tau peptide Cry-358DNITHVPGGGN368 was synthesized and used as an antigen to immunize C57BL/6 J mice for 4 times within two months. Antisera were pooled and the immunoreactivities to tauN368 and full-length hTau were tested by ELISA. Two mice with well serum immunization validated by Western blotting were selected, whose spleens were harvested for cell fusion. Positive hybridoma cells were selected by hypoxanthine-aminopterin-thymidin (HAT) medium, and the positive hybridoma cells were subcloned by limited dilution method to obtain monoclonal cell lines. During each subcloning, indirect ELISA screening was performed for 2–3 rounds to obtain positive monoclonal cell lines and then the antibody subtypes of the constructed cell lines were identified. Balb/c mice were injected with selected cell line, and ascitic fluid was purified for TauN368 antibody (titer > 1:64,000). All TauN368 Western blotting results were obtained using the newly generated antibody, while all TauN368 immunostaining results were obtained using the polyclonal rabbit anti-TauN368 antibody.

Western blotting

Mouse brains were removed, and the cortex and hippocampus were dissected on ice, respectively. Samples were homogenized with RIPA lysis buffer containing 50 mM Tris (pH 7.4), 150 mM NaCl, 1% Triton X-100, 1% sodium deoxycholate, and 0.1% SDS (#P0013B, Beyotime, Shanghai, China) mixed with a cocktail of protease and phosphatase inhibitors (Thermo Scientific, Waltham, MA), at a ratio of 10 μl/mg tissue. Then the tissue homogenates were centrifuged for 20 min at 12,000 ×g, resulting in RIPA-soluble and RIPA-insoluble parts. Protein concentration in the RIPA-soluble lysate was quantitated using BCA protein assay kit (Thermo Fisher), and equal amounts of proteins were loaded on SDS–PAGE gels. The RIPA-insoluble pellets were further dissolved in 90–120 μl of loading buffer containing 200 mM Tris–HCl pH 6.8, 8% SDS and 40% glycerol. Proteins were separated by SDS-PAGE (10%), transferred onto nitrocellulose membranes (Merck Millipore, Darmstadt, Germany) and then blocked with 5% bovine serum albumin (BSA). Membranes were incubated with primary and secondary antibodies (Additional file 1: Table S2), in sequence. Blots were visualized by an enhanced chemiluminescence substrate system (Santa Cruz Biotech, Santa Cruz, CA), imaged by an Odyssey Imaging System (LI-COR Biosciences, Lincoln, NE), and quantified by the ImageJ software. β-Actin was used as a loading control.

Immunostaining and quantification

Mice were anesthetized with 2% isoflurane (RWD Life Science, Shenzhen, China) one day after completion of the final behavioral trial, and perfused via the ventriculus sinister with 0.9% NaCl for 5 min followed by PBS containing 4% paraformaldehyde for 5 min. Brains were cryoprotected sequentially in 25% and 30% sucrose, for 2 days in total, and then cut into 30-μm sections using a cryostat microtome (CM1900, Leica, Wetzlar, Germany). For immunohistochemistry, free floating sections were immersed in 3% H2O2 in anhydrous methanol for 30 min, and non-specific sites were blocked with BSA for 30 min at room temperature. Brain slices were then incubated overnight at 4 °C with primary antibodies. Immunoreactions were developed using a DAB-staining kit (ZSGB-BIO, Bei**g, China). Images were taken by an automatic slice scanning system (SV120, Olympus, Tokyo, Japan) at 20 × magnification and analyzed with ImageJ software. Areas of various regions of the brain were measured.

For immunofluorescence, sections were thoroughly washed with PBST (PBS containing 0.1% Triton X-100) and incubated overnight with primary antibodies under 4 °C. After that, the sections underwent PBST washes for 15 min, followed by 1-h incubation with the secondary antibody at 37 °C, and finally counterstained with DAPI. Images were taken by an automatic slice scanning system (SV120, Olympus) and a two-photon laser-scanning confocal microscope (LSM 800, Zeiss, Oberkochen, Germany) at 20 × magnification, and analyzed with the ImageJ software. The areas of hTau368 staining in various regions, the mean intensity of NeuN staining in the CA1 pyramidal layer, as well as the numbers of cells positive for MAP2, GFAP, Iba1 or DCX staining were measured or counted in a single 20 × magnification view and averaged per 0.1 mm2. One section from each mouse and a total of four to six mice per group were analyzed.

Gallyas silver staining

Free-floating brain sections were washed with Tris-buffered saline (TBS) for 3 × 5 min, and then placed in 5% periodic acid followed by alkaline silver iodide solution and developer solution (#G1052, Servicebio, Wuhan, China). After washing with acetic acid and water, they were placed in 0.1% gold chloride, followed by sodium thiosulphate solution. The sections were placed in absolute ethanol for 3 × 5 min and in xylene for 2 × 5 min to become transparent. Images were taken by an automatic slice scanning system (SV120, Olympus, Tokyo, Japan) and analyzed at 100 × magnification.

Thioflavin S staining

Free-floating brain sections were washed with TBS for 3 × 5 min, and then incubated with 0.3% Thioflavin S (#1326-12-1, Sigma, Darmstadt, Germany) dissolved in 50% ethanol. The sections were then decolorized in 50% ethanol for 3 × 5 min, washed in TBS, and subsequently co-stained with DAPI for 10 min.

Sholl analysis

Fluorescent images of GFAP and Iba1 immunostaining were analyzed as reported previously [27]. Processes were traced by the plugin Sholl Analysis in ImageJ. The intersections of processes were counted within concentric circles at 10-μm intervals from the center of the soma. Ending radius and the sum of intersections were used to indicate the complexity of processes.

Electron microscopy

Synaptic density and neuronal axon width were determined by electron microscopy as described previously [28]. After deep anesthesia, mice were perfused transcardially with 2% glutaraldehyde and 3% paraformaldehyde in PBS. Hippocampal slices were post-fixed in cold 1% OsO4 for 1 h. Samples were prepared and examined using standard procedures. Ultrathin sections (90 nm) were stained with uranylacetate and lead acetate and viewed at 200 kV in a Tecnai electron microscope. The slices were from three mice in each group. Synaptic contacts were identified by the presence of synaptic vesicles and post-synaptic densities with high electron density.

Transcriptomic analysis

Hippocampal tissues were isolated on ice. The detailed procedures of total RNA extraction, mRNA library construction and quality assurance has been reported previously [29]. The differentially expressed genes (DEG) between vehicle-treated (n = 3) and Dox-treated hTau368 mice (n = 3) (FDR-adjusted P < 0.05, fold change > 1.5) were analyzed by DESeq2 (v1.4.5) [30]. Kyoto encyclopedia of genes and genomes (KEGG) enrichment and the relationship network of KEGG pathways of DEGs were analyzed by Dr. Tom software (BGI, Shenzhen, China). Gene network analysis was performed using the STRING database (https://string-db.org/).

Novel location recognition test

Mice were handled and habituated before tests. On day 1, each mouse was placed in the center of a plastic box in which two identical objects (A and B) were located in two corners. The mouse was allowed 5 min to explore freely. After 24 h, the mouse was placed back to the box with object A in the same corner while object B placed in a new location. Again, each mouse was allowed 5 min to explore. The time the mouse spent exploring objects A and B was recorded as TA and TB, respectively. Exploration was identified by a video tracking system (Anymaze Technology SA, Stoelting Co., IL) once the mouse head stayed close (< 3 cm) to either object. Mice with TA or TB less than 2 s were excluded from analysis. The discrimination index was calculated as (TB−TA)/(TB + TA). A higher discrimination index indicates better spatial memory.

Morris water maze test

Mice were kept in the test room for 24 h before tests. In the learning phase, mice were trained in the maze to find the hidden platform. The learning phase consisted of 3 trials per day with an interval of 30 min, from 14:00 to 17:00, for 5 consecutive days. In each trial, a mouse was placed in one of the three quadrants without the platform, facing the wall of the pool. If the mouse found the hidden platform within 60 s, another 15 s was left for learning consolidation. If the mouse did not find the platform within 60 s, it was guided to the platform and allowed to stay on the platform for 15 s. The time to find the platform during the 5-day training was recorded as the escape latency. On day 6, a testing trial was performed. The hidden platform was removed and each mouse was placed in the quadrant opposite to the target quadrant. The time/distance and trajectory of each mouse traveling in the pool were recorded and analyzed by a video tracking system (Chengdu Taimeng Software Co., Ltd, China). Mice disabled in eyes or limbs were excluded from analysis.

Open field test

Mice were handled for 1 day, and placed in the test room the day before the behavioral test to get acclimated to the environment. The open field apparatus was a 60 × 60 × 50 cm3 white plastic box, with the floor divided virtually into 16 equal squares in the monitoring system, including a central field (the central 4 square regions) and 12 periphery fields. Each mouse was allowed to explore freely in the box for 5 min. The time and distance each mouse travelled in different zones were recorded and analyzed by the ANY-maze video tracking system (Stoelting Co., WoodDale, IL).

Elevated-plus maze test

The elevated-plus maze consisted of two enclosed arms (65 × 5 × 20 cm3) and two open arms (65 × 5 cm2). The apparatus was elevated to 50 cm above the floor. Each mouse was placed in the center of the maze, facing the open arm opposite to the experimenter, and allowed to explore freely for 5 min. The time and place mice traveled in the maze were recorded and analyzed using a video tracking system (Chengdu Taimeng Software Co., Ltd, China).

Statistical analyses

All data were analyzed and plotted using GraphPad Prism 8 (La Jolla, CA). Comparisons between two groups were made by two-tailed unpaired Student’s t-tests, and comparisons between multiple groups were conducted with one-way, two-way or repeated measures ANOVA followed by post-hoc tests for multiple comparisons. P < 0.05 was considered statistically significant. All experiments and analysis were performed in a blind manner. All values are shown as mean ± SEM or min-median-max unless otherwise specified.

Results

Generation of the hTau368 transgenic mice

To design a chimeric transgenic vector, the human MAPT gene encoding the hTau368 fragment was placed downstream the tetracycline-responsive element (TRE) promoter, which joined to assemble a tet-on system with a second module expressing the reverse tetracycline-controlled transactivator (rtTA) [31, 32] under the control of the neuron-specific Eno2 (also known as NSE) promoter [33, 34]. These two parts were reversely linked through poly-adenine, and chicken beta-globin insulator was added at both 3′ and 5′ ends to prevent position effects (Fig. 1a). The transgenic vector was constructed and subsequently confirmed by PCR using flanking and insert-specific primers and with restriction analyses. Vectors were transferred into fertilized eggs from C57BL/6 J mice through microinjections, and then engrafted onto the uterus wall of pregnant mice. Successful vector transfection was identified through genoty** in a F0 offspring. Whole-genome sequencing showed that the transgenic vector was randomly inserted into the intron between exons 9 and 10 of gene Mindy 3 on Chromosome 2. This transgenic mouse was subsequently used for the initial breeding (Additional file 1: Fig. S1).

Fig. 1
figure 1

Predominant hTau expression in the hippocampus of dox-administered hTau368 mice. a Strategy to express hTau1-368 fragment (hTau368) in transgenic mice. The hTau368 expression was controlled by a tet-on system in combination with the neuronal specific Eno2 promoter. In the presence of doxycycline (dox-on), reverse tetracycline transactivator (rtTA) binds to the tetracycline-responsive element (TRE) to initiate hTau368 expression. In the absence of doxycycline (dox-off), hTau368 expression stops, as rtTA cannot bind to TRE. b–d Relatively prominent immunofluorescent intensity of hTau368 in the hippocampus following 2-month dox treatment. Representative sagittal (b) and coronal (c) images are shown. One-way ANOVA followed by Tukey's multiple comparisons tests, *** P < 0.001, n = 4 sections from 4 mice. Data were normalized to the mean value in the hippocampus. e, f Western blotting showed relatively prominent human tau expression in the hippocampus of dox-treated hemizygous hTau368 mice. One-way ANOVA followed by Tukey’s multiple comparisons tests, *** P < 0.001, compared with the hippocampus; n = 3 mice in each group. g, h hTau368 distribution in both neuronal soma and neurites in multiple hippocampal subregions. One-way ANOVA followed by Tukey's multiple comparisons tests, *P < 0.05, compared with CA2; n = 4 sections from 4 mice. Data were normalized to the mean value in CA2. i, j hTau368 mice (hemizygous) showed time-dependent increase of hTau expression in the hippocampus following dox administration. One-way ANOVA followed by Tukey's multiple comparisons tests, *P < 0.05, ***P < 0.001; n = 3 mice in each group, sacrificed at 15–16 months old. Data were normalized to the mean value of the 1-month Dox-treatment group. k, l The hTau level in the hippocampus of hTau368 mice (hemizygous) gradually decreased during 1–3 months after dox retraction (dox-off). n = 4 mice in each group

Full size image

We next examined the distribution of hTau368 in the brain following dox administration in drinking water (2 mg/l) for 2 months, starting at 2 months of age. Following the 2-month dox treatment, we detected prominent hTau expression in the hippocampus of hemizygous hTau368 mice (Fig. 1b–h), the best-recognized area responsible for cognitive impairments in AD. By contrast, only weak tau signal was detected in other areas of the central nerves system, such as cerebral cortex, forebrain, cerebellum, brainstem and spinal cord (Fig. 1b–h and Additional file 1: Fig. S2). Remarkable hTau aggregation from neurites to soma was observed in hippocampal subregions, including DG, CA3, CA2, CA1 and subiculum (Fig. 1g, h). Also, we found mild hTau368 expression in the entorhinal-piriform cortex (EC and Pir) and amygdala (Fig. 1c and Additional file 1: Fig. S2). Hippocampus, entorhinal cortex and amygdala are vulnerable regions to AD, while cerebellum, brainstem and spinal cord are irrelevant brain areas [35]. These results suggest that in this model, hTau368 is expressed in brain regions susceptible to AD.

Then we focused on hippocampus due to the highest hTau expression level in this region. We found that hTau was accumulated in a time-dependent manner, and dox treatment for one month was enough to induce overt hTau expression in hemizygous hTau368 mice (Fig. 1i, j). Unexpectedly, the hTau expression probed by Tau368 or HT7 antibody (which specifically reacts with hTau proteins) progressively diminished and almost disappeared at 3 months after dox retraction (Fig. 1k, l). We also observed that accumulation of the phosphorylated tau at AT8 (pThr202 & pThr205) epitope was remarkably relieved following dox-off for 3 months, especially in the apical dendrites of CA1 pyramidal neurons and DG (Additional file 1: Fig. S3). These results together indicate that the hTau368 transgenic line is an easy-to-use, inducible and reversible model for studying tau pathologies in AD and related tauopathies, especially when we are focusing on the hippocampus.

Phosphorylated tau accumulation in the hippocampus of dox-treated hTau368 mice

Hyperphosphorylation of tau is the major driver of NFT formation [4]. Over 30 phosphorylation sites have been reported to correlate with tau pathology in AD [36, 55], and peptides designed against this 306VQIVYK311 fragment are capable of inhibiting tau fibril formation [56, 57]. In postmortem brains of AD patients, tau fragments ending at the N368 site are present in NFTs [58]. These data together strongly support the feasibility of tau 1–368 fragment in mimicking the AD-like tau model.

Unexpectedly, we found here that the tau-associated pathologies and cognitive impairment induced by 2-month dox treatment in hTau368 mice gradually relieved following dox withdrawn for 3 months. This is consistent with a previous report that in a TAU62 transgenic line with dox-dependent 3R tau 151–421 expression, the nerve cell dysfunction and severe paralysis generally recovered when the expression of tau was halted [51]. In another line rTg4510 which uses a tet-off system to control the expression of mutant P301L hTau, tau-associated pathologies like NFTs, neuronal loss, forebrain atrophy and memory impairments were also ameliorated following the cessation of hTau expression [16]. The mechanism for this phenomenon may involve activation of a compensatory system to remove pathological tau, such as activation of the proteasome or lysosomal proteolysis system, and modulation of tau-associated protein kinases and phosphatases which regulate tau phosphorylation and thus indirectly regulate tau degradation. Simultaneously, synaptic remodeling and loss of hippocampal neurons ceased when hTau had been eliminated, which confirmed the toxic role of hTau368. It remains to be determined whether the gradual and autonomic alleviation of tau pathology will still happen when dox-on duration persists for a much longer time, like 6–9 months; the underlying mechanisms warrant further investigations to understand the etiology and unravel the drug targets of AD from the perspective of tauopathy. Additionally, we only presented the results of dox-induction in young animals. As AD is an ageing-associated disorder, the dox-induced tau pathology in older animals needs to be studied. We anticipate that the time to the appearance of similar pathologies and behavioral deficits caused by dox induction should be shorter in older mice.

Moreover, in the brains of the elderly and AD patients, there should be more tau x–368 fragments, which are all cleaved at N368 but at different sites from the C-terminal on the other end. It remains to be defined the content of each fragment in human AD brains, especially the longest 1–368 fragment. We designed the tau368 mice here to express hTau 1–368 since it showed relatively higher cytotoxicity than many other tau fragments [25], while it is also possible for the hTau1-368 fragment to be further cleaved by murine AEP or other endogenous enzymes to generate smaller-size and toxic tau species, like what happens in human AD brains. In addition to the hTau368 mice, other transgenic mouse lines expressing truncated hTau, such as tau159–391 which is also found in AD patients [59], might also be good tools to study tau-associated pathologies in AD.

In previously reported amyloid models, such as PDAPP [60], APPswe/PSEN1dE9 (JAX034832) and 5 × FAD (JAX034848), the age-dependent increase of Aβ pathology is significant while tau pathologies are minor or appear at a relatively old age [36, 61]. As for tau models such as Tau3R0N (hMAPT3R0N, JAX003741), P301L-Tau0N4R (rTg4510, JAX015815 and 016198) and P301S-Tau1N4R (PS19, JAX008169), no significant amyloid deposition is reported [10, 36,

Availability of data and materials

All data provided in this paper are available from the leading contact, Prof Jian-Zhi Wang upon reasonable request.

Change history

Abbreviations

EC:

Entorhinal cortex

Pir:

Piriform cortex

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Acknowledgements

We thank Prof. Keqiang Ye for providing the TauN368 antibody, and **gfen Su and Yue ** Liu & Jian-Zhi Wang

  • Department of Neurology, Renmin Hospital of Wuhan University, Wuhan, 430030, China

    Zhentao Zhang

  • Key Laboratory of Ministry of Education for Neurological Disorders, Department of Endocrine, Liyuan Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430077, China

    Yao Zhang

  • Neuroscience Research Institute and Department of Neurobiology, School of Basic Medical Sciences, Peking University, Bei**g, China

    Jie Zheng

  • Key Laboratory for Neuroscience, Ministry of Education/National Health Commission, Peking University, Bei**g, 100083, China

    Jie Zheng

  • Co-Innovation Center of Neuroregeneration, Nantong University, Nantong, 226000, China

    Jian-Zhi Wang

    • Authors
    1. Yang Gao
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    2. Yuying Wang
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    3. Huiyang Lei
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    Contributions

    J.Z.W., J.Z., Y.Z. and Y.G. designed this research; Y.G., Y.W., H.L., Z.X., S.L., H.Y., X.J., Z.X. and Y.Z. performed the experiments. Z.Z. provided advices on this research. Y.G, Y. W., Y.Z and Z.J. performed statistical analysis and data interpretation. Y.G., J. Z. and J.Z.W. wrote the manuscript. All authors read and approved the final manuscript. J.Z.W. is the supervisor of this work who has full access to all the data in the study and takes responsibility for the data integrity and accuracy.

    Corresponding authors

    Correspondence to Yang Gao, Yao Zhang, Jie Zheng or Jian-Zhi Wang.

    Ethics declarations

    Ethics approval and consent to participate

    All animal studies had complied with all relevant ethical regulations for the animal testing and research, and were approved by institutional guidelines and the Animal Care and Use Committee of Tongji Medical College, Huazhong University of Science and Technology.

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    Not applicable.

    Competing interests

    The authors declare that they have no competing interests.

    Additional information

    The original online version of this article was revised: Figure 2 has been corrected.

    Supplementary Information

    Additional file 1: Fig. S1

    . Generation and genomic identification of hTau368 mice. Fig. S2. hTau368 had predominantly expression in hippocampus, slightly in other regions. Fig. S3. Reversible tau phosphorylation in hTau368 mice following dox-off. Fig. S4. Dox treatment increased hTau in the pan-cortex of hTau368 mice. Fig. S5. Dox-treated hTau368 mice showed enhanced Gallyas silver staining in DG granular cells, although the staining intensity was much slighter than that detected in the brain slice of AD patients. Fig. S6. Dox-treated hTau368 mice did not show amyloid deposition. Fig. S7. Dox treatment upregulated GSK-3β activity in the hippocampus of hTau368 mice. Fig. S8. Enhanced gliosis in entorhinal-piriform cortex of dox-treated hTau368 mice. Fig. S9. Dox treatment showed limited effect on glia activation in wild-type mice. Fig. S10. Reduction of tau correlates with increased synapse-associated proteins in hTau368 mice. Fig. S11. The loss of hippocampal neurons ceased when dox was retracted for hTau368 mice. Fig. S12. Dox-treated hTau368 mice tended to exhibit increased locomotor activities. Fig. S13. hTau368 mice showed no gender difference in tauopathy and cognitive behaviors. Table S1. Primers used for the identification of hTau368 mice. Table S2. Antibodies used in this study.

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    Gao, Y., Wang, Y., Lei, H. et al. A novel transgenic mouse line with hippocampus-dominant and inducible expression of truncated human tau. Transl Neurodegener 12, 51 (2023). https://doi.org/10.1186/s40035-023-00379-5

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