Integrative analysis of γδT cells and dietary factors reveals predictive values for autism spectrum disorder in children.

BACKGROUND: Autism spectrum disorder (ASD) includes a range of multifactorial neurodevelopmental disabilities characterized by a variable set of neuropsychiatric symptoms. Immunological abnormalities have been considered to play important roles in the pathogenesis of ASD, but it is still unknown which abnormalities are more prominent. METHODS: A total of 105 children with ASD and 105 age and gender-matched typically developing (TD) children were recruited. An eating and mealtime behavior questionnaire, dietary habits, and the Bristol Stool Scale were investigated. The immune cell profiles in peripheral blood were analyzed by flow cytometry, and cytokines (IFN-γ, IL-8, IL-10, IL-17A, and TNF-α) in plasma were examined by Luminex assay. The obtained results were further validated using an external validation cohort including 82 children with ASD and 51 TD children. RESULTS: Compared to TD children, children with ASD had significant eating and mealtime behavioral changes and gastrointestinal symptoms characterized by increased food fussiness and emotional eating, decreased fruit and vegetable consumption, and increased stool astriction. The proportion of γδT cells was significantly higher in children with ASD than TD children (ß: 0.156; 95% CI: 0.888 âˆ¼ 2.135, p < 0.001) even after adjusting for gender, eating and mealtime behaviors, and dietary habits. In addition, the increased γδT cells were evident in all age groups (age < 48 months: ß: 0.288; 95% CI: 0.420 âˆ¼ 4.899, p = 0.020; age ≥ 48 months: ß: 0.458; 95% CI: 0.694 âˆ¼ 9.352, p = 0.024), as well as in boys (ß: 0.174; 95% CI: 0.834 âˆ¼ 2.625, p < 0.001) but not in girls. These findings were also confirmed by an external validation cohort. Furthermore, IL-17, but not IFN-γ, secretion by the circulating γδT cells was increased in ASD children. Machine learning revealed that the area under the curve in nomogram plots for increased γδT cells combined with eating behavior/dietary factors was 0.905, which held true in both boys and girls and in all the age groups of ASD children. The decision curves showed that children can receive significantly higher diagnostic benefit within the threshold probability range from 0 to 1.0 in the nomogram model. CONCLUSIONS: Children with ASD present with divergent eating and mealtime behaviors and dietary habits as well as gastrointestinal symptoms. In peripheral blood, γδT cells but not αßT cells are associated with ASD. The increased γδT cells combined with eating and mealtime behavior/dietary factors have a high value for assisting in the diagnosis of ASD.

Tmem30a Plays Critical Roles in Ensuring the Survival of Hematopoietic Cells and Leukemia Cells in Mice.

The fundamental structure of eukaryotic cell plasma membrane is the phospholipid bilayer, which contains four major phospholipids. These phospholipids are asymmetrically distributed between the outer and inner leaflets. P4-ATPase flippase complexes play essential roles in ensuring this asymmetry. We found that conditional deletion of Tmem30a, the ß subunit of P4-ATPase flippase complex, caused pancytopenia in mice. Tmem30a deficiency resulted in depletion of lineage-committed blood cells in the peripheral blood, spleen, and bone marrow. Ablation of Tmem30a also caused the depletion of hematopoietic stem cells (HSCs). HSC RNA sequencing results revealed that multiple biological processes and signal pathways were involved in the event, including mammalian target of rapamycin signaling, genes for HSC stemness, and genes responding to interferons. Our results also revealed that targeting Tmem30a signaling had therapeutic utility in BCR/ABL1-induced chronic myeloid leukemia.

Atp8a1 deletion increases the proliferative activity of hematopoietic stem cells by impairing PTEN function.

PURPOSE: The eukaryotic cell plasma membrane contains several asymmetrically distributed phospholipids, which is maintained by the P4-ATPase flippase complex. Herein, we demonstrated the biological effects and mechanisms of asymmetrical loss in hematopoietic stem cells (HSCs). METHODS: An Atp8a1 knockout mouse model was employed, from which the HSC (long-term HSCs and short-term HSCs) population was analyzed to assess their abundance and function. Additionally, competitive bone marrow transplantation and 5-FU stress assays were performed. RNA sequencing was performed on Hematopoietic Stem and Progenitor Cells, and DNA damage was assayed using immunofluorescence staining and comet electrophoresis. The protein abundance for members of key signaling pathways was confirmed using western blotting. RESULTS: Atp8a1 deletion resulted in slight hyperleukocytosis, associated with the high proliferation of HSCs and BCR/ABL1 transformed leukemia stem cells (LSCs). Atp8a1 deletion increased the repopulation capability of HSCs with a competitive advantage in reconstitution assay. HSCs without Atp8a1 were more sensitive to 5-FU-induced apoptosis. Moreover, Atp8a1 deletion prevented HSC DNA damage and facilitated DNA repair processes. Genes involved in PI3K-AKT-mTORC1, DNA repair, and AP-1 complex signaling were enriched and elevated in HSCs with Atp8a1 deletion. Furthermore, Atp8a1 deletion caused decreased PTEN protein levels, resulting in the activation of PI3K-AKT-mTORC1 signaling, further increasing the activity of JNK/AP-1 signaling and YAP1 phosphorylation. CONCLUSION: We identified the role of Atp8a1 on hematopoiesis and HSCs. Atp8a1 deletion resulted in the loss of phosphatidylserine asymmetry and intracellular signal transduction chaos.

Single-cell RNA transcriptome landscape of murine liver following systemic administration of mesoporous silica nanoparticles.

Due to the unique physicochemical properties, mesoporous silica nanoparticles (MONs) have been widely utilized in biomedical fields for drug delivery, gene therapy, disease diagnosis and imaging. With the extensive applications and large-scale production of MONs, the potential effects of MONs on human health are gaining increased attention. To better understand the cellular and molecular mechanisms underlying the effects of MONs on the mouse liver, we profiled the transcriptome of 63,783 single cells from mouse livers following weekly intravenous administration of MONs for 2 weeks. The results showed that the proportion of endothelial cells and CD4+ T cells was increased, whereas that of Kupffer cells was decreased, in a dose-dependent manner after MONs treatment in the mouse liver. We also observed that the proportion of inflammation-related Kupffer cell subtype and wound healing-related hepatocyte subtype were elevated, but the number of hepatocytes with detoxification characteristics was reduced after MONs treatment. The cell-cell communication network revealed that there was more crosstalk between cholangiocytes and Kupffer cells, liver capsular macrophages, hepatic stellate cells, and endothelial cells following MONs treatment. Furthermore, we identified key ligand-receptor pairs between crucial subtypes after MONs treatment that are known to promote liver fibrosis. Collectively, our study explored the effects of MONs on mouse liver at a single-cell level and provides comprehensive information on the potential hepatotoxicity of MONs.

Single-cell transcriptomic dissection of the cellular and molecular events underlying the triclosan-induced liver fibrosis in mice.

BACKGROUND: Triclosan [5-chloro-2-(2,4-dichlorophenoxy) phenol, TCS], a common antimicrobial additive in many personal care and health care products, is frequently detected in human blood and urine. Therefore, it has been considered an emerging and potentially toxic pollutant in recent years. Long-term exposure to TCS has been suggested to exert endocrine disruption effects, and promote liver fibrogenesis and tumorigenesis. This study was aimed at clarifying the underlying cellular and molecular mechanisms of hepatotoxicity effect of TCS at the initiation stage. METHODS: C57BL/6 mice were exposed to different dosages of TCS for 2 weeks and the organ toxicity was evaluated by various measurements including complete blood count, histological analysis and TCS quantification. Single cell RNA sequencing (scRNA-seq) was then carried out on TCS- or mock-treated mouse livers to delineate the TCS-induced hepatotoxicity. The acquired single-cell transcriptomic data were analyzed from different aspects including differential gene expression, transcription factor (TF) regulatory network, pseudotime trajectory, and cellular communication, to systematically dissect the molecular and cellular events after TCS exposure. To verify the TCS-induced liver fibrosis, the expression levels of key fibrogenic proteins were examined by Western blotting, immunofluorescence, Masson's trichrome and Sirius red staining. In addition, normal hepatocyte cell MIHA and hepatic stellate cell LX-2 were used as in vitro cell models to experimentally validate the effects of TCS by immunological, proteomic and metabolomic technologies. RESULTS: We established a relatively short term TCS exposure murine model and found the TCS mainly accumulated in the liver. The scRNA-seq performed on the livers of the TCS-treated and control group profiled the gene expressions of > 76,000 cells belonging to 13 major cell types. Among these types, hepatocytes and hepatic stellate cells (HSCs) were significantly increased in TCS-treated group. We found that TCS promoted fibrosis-associated proliferation of hepatocytes, in which Gata2 and Mef2c are the key driving TFs. Our data also suggested that TCS induced the proliferation and activation of HSCs, which was experimentally verified in both liver tissue and cell model. In addition, other changes including the dysfunction and capillarization of endothelial cells, an increase of fibrotic characteristics in B plasma cells, and M2 phenotype-skewing of macrophage cells, were also deduced from the scRNA-seq analysis, and these changes are likely to contribute to the progression of liver fibrosis. Lastly, the key differential ligand-receptor pairs involved in cellular communications were identified and we confirmed the role of GAS6_AXL interaction-mediated cellular communication in promoting liver fibrosis. CONCLUSIONS: TCS modulates the cellular activities and fates of several specific cell types (including hepatocytes, HSCs, endothelial cells, B cells, Kupffer cells and liver capsular macrophages) in the liver, and regulates the ligand-receptor interactions between these cells, thereby promoting the proliferation and activation of HSCs, leading to liver fibrosis. Overall, we provide the first comprehensive single-cell atlas of mouse livers in response to TCS and delineate the key cellular and molecular processes involved in TCS-induced hepatotoxicity and fibrosis.

Targeting glutaminase1 and synergizing with clinical drugs achieved more promising antitumor activity on multiple myeloma.

Multiple myeloma (MM) pathogenesis remains incompletely understood and biomarkers predicting treatment response still remain lacking. Here we describe the rational mechanisms of combining targeting glautaminase1 (GLS1) with other chemo-reagents for MM treatment. Gls1 is highly expressed cMYC/KRAS12V-drived plasmacytoma (PCT) cells. Down-regulation of Gls1 with miRNAi in cMYC/KRAS12V-expressing BaF3 cells prevented them from growing independence of interleukin 3 (IL3). By using our cMYC/KRAS12V-transduced adoptive plasmacytoma mouse model, we found that Gls1 is involved in PCT pathogenesis. Down-regulation of Gls1 significantly prolonged the survival of PCT recipients. Knockdown of Gls1 increased the expression of Cdkn1a and Cdkn1b and decreased the expression of some critical oncogenes for cancer cell survival, such as c-Myc, Cdk4, and NfκB, as well as some genes which are essential for MM cell survival, such as Irf4, Prdm1, Csnk1α1, and Rassf5. Combination of Gls1 inhibition with LBH589, Bortezomib, or Lenalidomide significantly impaired tumor growth in a MM xenograft mouse model. Our data strongly suggest that Gls1 plays an important role for MM pathogenesis and that combination of GLS1 inhibitor with other MM therapy agents could benefit to MM patients.