Journal: Advanced Science

Article Title: Targeting Microglial CD49a Inhibits Neuroinflammation and Demonstrates Therapeutic Potential for Parkinson's Disease

doi: 10.1002/advs.202515138

Figure Lengend Snippet: Microglial transcriptome analysis revealed a protective role of Itga1 knockdown in a Parkinson's disease (PD) mouse model. (A) A schematic diagram of microglial isolation, RNA extraction, RNA‐Seq, and bioinformatic analysis. (B) Venn diagram showing the CDEGs (277) between the “shCon/MPTP vs shCon/Saline” groups (1446) and “shITGA1/MPTP vs shCon/MPTP” groups (526). (C) Hierarchical clustered heatmap of gene expression profiles for the common differentially expressed genes (CDEGs). (D,E) Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway and Gene Ontology Biological Processes (GO‐BP) analysis of the CDEGs. Green arrows point to neurodegenerative KEGG pathways; red arrows point to inflammatory GO‐BP. (F) Representative immunofluorescence images depicting morphological analysis (Outline and Skeleton) of IBA1 + microglia in the SNc. Microglia borders are marked by dashed lines. Scale bar: 10 µm. (G–J) Quantification of the soma's area, number of endpoints, number of branches, and maximal branch length in microglia within the SNc ( n = 10). (K) Quantification of IL‐1β, IL‐6, IL‐18, and TNF‐α concentration levels in mouse striatal homogenate detected by ELISA ( n = 3). Data are presented as means ± SEM with one‐way and two‐way ANOVA. * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001.

Article Snippet: Mouse IL‐1β (EK0394), IL‐6 (EK0411), IL‐18 (EK0433), and TNF‐α (EK0527) Enzyme‐Linked Immunosorbent Assay (ELISA) Kits were acquired from Boster Biotechnology.

Techniques: Knockdown, Isolation, RNA Extraction, RNA Sequencing, Saline, Gene Expression, Immunofluorescence, Concentration Assay, Enzyme-linked Immunosorbent Assay

Journal: Advanced Science

Article Title: Targeting Microglial CD49a Inhibits Neuroinflammation and Demonstrates Therapeutic Potential for Parkinson's Disease

doi: 10.1002/advs.202515138

Figure Lengend Snippet: Pharmacological inhibition of CD49a relieves neuroinflammation and improves motor impairments in a Parkinson's disease (PD) mouse model. (A) Molecular docking analysis of obtustatin (PDB: 1MPZ) with integrin α1β1 (PDB: 1QC5) (left) and the potential interaction binding site (right). Obt, obtustatin; Green part, integrin α1β1 (Chain A); blue part, integrin α1β1 (Chain B); magenta part, obtustatin. (B) Experimental design involving obtustatin treatment (i.n.; 5 mg/kg) in an 1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine (MPTP) induced chronic PD mouse model (i.p.; 25 mg/kg). Obtustatin and MPTP were administered twice weekly for 5 weeks. Obtustatin was administered in the morning and MPTP in the afternoon, with an interval of 6 h. Behavioral tests were performed in Week 6. The experimental animals were sacrificed in Week 7. (C) Travelled trace of mice in the open field test (OFT). (D,E) Quantification of the total distance, distance in center, and not moving time in the OFT as well as quantification of the grip strength test, rotarod test, and pole‐climbing test ( n = 10). (F,L) Representative immunohistochemical staining images and quantification of TH + density in the striatum. Scale bar: 200 µm. (G,M) Representative immunohistochemical staining images and quantification of TH + neurons in the SNc ( n = 3). Borders of the SNc are shown by ellipses. Scale bar: 100 µm. (H,N) Representative Nissl staining images and quantification in the SNc ( n = 3). Scale bar: 100 µm. Borders of the SNc are shown by ellipses. (I,P) Ultrastructural images and quantification of mitochondria in the SNc ( n = 5). Scale bar: 200 nm. Black arrows represent mitochondria. The green, yellow, and red in the statistical chart represent normal, damaged, and degenerated mitochondria, respectively. (J) Representative immunofluorescence images depicting morphological analysis (Skeleton) of IBA1 + microglia in the SNc of mice. Microglia borders are marked by dashed lines. Scale bar: 10 µm. (K,O) Representative blots and quantification of TH expression levels in the SN ( n = 3). (Q–T) Quantification of soma's area, number of endpoints, number of branches, and maximal branch length in SNc microglia. ( n = 10). (U) Quantification of dopamine concentrations in striatal tissue homogenates measured using ELISA ( n = 3). (V) Quantification of IL‐1β, IL‐6, IL‐18, and TNF‐α concentration levels in mouse striatum homogenate detected by ELISA ( n = 3). Data are presented as means ± SEM with t ‐test and one‐way ANOVA. * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001.

Article Snippet: Mouse IL‐1β (EK0394), IL‐6 (EK0411), IL‐18 (EK0433), and TNF‐α (EK0527) Enzyme‐Linked Immunosorbent Assay (ELISA) Kits were acquired from Boster Biotechnology.

Techniques: Inhibition, Binding Assay, Immunohistochemical staining, Staining, Immunofluorescence, Expressing, Enzyme-linked Immunosorbent Assay, Concentration Assay

Journal: Advanced Science

Article Title: Loss of SOCS1 in Donor T Cells Exacerbates Intestinal GVHD by Driving a Chemokine‐Dependent Pro‐Inflammatory Immune Microenvironment

doi: 10.1002/advs.202513735

Figure Lengend Snippet: Socs1 deficiency in T cells drives effector differentiation and enhances inflammatory responses in CD8 + T cells. (A) Experimental schematic. Splenocytes from WT (littermate control; Socs1 fl/fl ) and cKO (LckCre‐ Socs1 fl/fl ) mice were isolated and sorted by FACS for CD45 + cells and subjected to single‐cell RNA sequencing (scRNA‐seq, n = 5 per group). Alternatively, CD8 + T cells from the spleen of WT and cKO mice were sorted by FACS and subjected to bulk RNA‐seq, ATAC‐seq, and CUT&Tag analyses. (B) UMAP plot of 105 040 single cells from CD45 + splenocytes colored by annotated immune cell subsets. (C) UMAP visualization of CD45 + splenocytes, split by origin. (D) Comparison of the proportions of celltypes between WT and cKO groups. (E) UMAP plot of 19 490 T cells extracted from Figure and colored by annotated T cell subsets. (F) UMAP visualization of T cells, split by origin. (G) Heatmap of Ro/e (Ratio of observed to expected) scores for T cell subtypes in WT and cKO mice. The scores, calculated from scRNA‐seq cell counts, indicate the relative enrichment (red, Ro/e > 1) or depletion (white/light orange, Ro/e < 1) of each population within each genotype. Numerical values are presented alongside a semi‐quantitative summary. (H) UMAP plot of 9159 CD8 + T cells extracted from Figure and colored by annotated T cell subsets. (I) Comparison of the proportions of indicated CD8 + T cell clusters between WT and cKO groups. (J) Representative flow cytometry plots and frequencies of naive T cells (Tn; CD44 − CD62L + ), central memory (Tcm; CD44 + CD62L + ), and effector memory (Tem; CD44 + CD62L − ) in CD8 + T cells from peripheral blood (PB, left panel) and spleen (SP, right panel) (n = 5 per group). (K) Bar plots showing the expression of perforin, GZMB, TNF‐α, IFN‐γ, IL‐2, and CD107a in CD8 + T cells from WT and cKO mice, as measured by flow cytometry (n=5 per group). (L) UMAP visualization of integrated T‐cell transcriptomes from WT (left, 25 304 cells) and cKO (right, 21 588 cells) groups. Each point represents a single cell, colored by the frequency of its corresponding TCR clonotype, highlighting clonally expanded cells. (M) Quantification of overall TCR repertoire diversity. The Gini index (top) and Shannon entropy (bottom) were calculated for the entire T‐cell population from each mouse. (N) Distribution and clonal size of T cells across identified subsets. Barplot showing the absolute cell counts (left panels) and the clonal size composition (right panels) for each T‐cell subset from WT and cKO mice. Data represent one experiment out of two independent experiments. P values were determined using two‐sided Wilcoxon rank‐sum test (D, I, M) or unpaired two‐tailed Student's t‐test (J‐K). Data represent mean ± SEM (D, I, M) or mean ± SD (J‐K). ∗ p <.05, ∗∗ p <.01 and ∗∗∗∗ p <.0001.

Article Snippet: The following kits were used: IL‐1β (Proteintech, KE10003), IL‐6 (Proteintech, KE10007), and TNF‐α (Proteintech, KE10002).

Techniques: Control, Isolation, Single Cell, RNA Sequencing, Comparison, Flow Cytometry, Expressing, Two Tailed Test

Journal: Advanced Science

Article Title: Loss of SOCS1 in Donor T Cells Exacerbates Intestinal GVHD by Driving a Chemokine‐Dependent Pro‐Inflammatory Immune Microenvironment

doi: 10.1002/advs.202513735

Figure Lengend Snippet: Evolution of small intestinal immune cell composition following transplantation of Socs1 cKO CD8 + T cells. (A) Experimental schematic. Lethally irradiated BALB/c recipient mice were transplanted with splenic T cells from WT mice (WT group) or cKO mice (cKO group), along with 5 × 10 6 TCD‐BM cells from WT mice. Survival was monitored daily. Body weight and GVHD score were assessed every five days. Immune cells in PB and small intestine from WT and cKO groups were assessed on Day 7, Day 16, and Day 24 by flow cytometry. (B) Survival analysis of recipients transplanted with 1 × 10 6 , 2 × 10 6, or 3 × 10 6 splenic T cells from WT or cKO mice (n = 10 mice per group). A control group received TCD‐BM only. Median survival times for cKO groups were 36 (1 × 10 6 ), 33 (2 × 10 6 ), and 26 days (3 × 10 6 ), respectively. In the corresponding WT groups, 9/10 mice in the 1 × 10 6 group survived to the end of the observation period, with median survival times of 35 days (2 × 10 6 ), and 42.5 days (3 × 10 6 ). Data were pooled from two independent experiments (n=10 mice/group). (C‐D) GVHD score (C) and body weight changes (D) in WT and cKO recipients transplanted with 1 × 10 6 splenic T cells. (E) Proportions of CD8 + T cells and monocytes in PB, and proportions of CD8 + T cells in the IEL and LP on Day 7, 16, and 24 post‐transplantation. (F) Boxplots showing the expression of TNF‐α, IFN‐γ, perforin, and CD107a in CD8 + T cells in IEL by flow cytometry on Day 7 and 16 post‐transplantation. (G) Experimental schematic of scRNA‐seq. Lymphocytes from IEL and LP were isolated and collected from 3‐4 mice, followed by FACS sorting for CD45 + donor‐derived (H‐2 b+ H‐2 d− ) cells respectively. Sorted cells from IEL and LP were mixed in a 1:2 ratio and subsequently subjected to scRNA‐seq. (H‐I) UMAP plot of all immune cells colored by samples (H) or annotated subsets (I). (J) UMAP plot showing annotated CD8 + T cell subsets (left), with barplot illustrating the proportion of each annotated subset in recipient intestines at indicated time points between WT and cKO groups (right). (K) Violin plots comparing the expression of cytotoxic molecules and cytokine receptor genes in CD8 + T cells on Day 26 post‐transplantation between WT and cKO groups. (L) UMAP plot showing annotated myeloid cell subsets (left), with barplot illustrating the proportion of each annotated subset in recipient intestines at indicated time points between WT and cKO groups (right). (M) Violin plots comparing expression of M1‐ and M2‐associated marker genes in myeloid cells on Day 26 post‐transplantation between WT and cKO groups. Data represent three independent experiments. P values were determined using chi‐squared test (B) or unpaired two‐tailed Student's t‐test (E, F) or two‐sided Wilcoxon rank‐sum test (K, M). Differences in GVHD scores and body weight between the WT and cKO groups at each time point were analyzed using multiple unpaired two‐tailed Student's t ‐tests (C, D). Data represent mean ± SEM (C‐E). ∗ p <.05, ∗∗ p <.01, ∗∗∗ p <.001 and ∗∗∗∗ p <.0001.

Article Snippet: The following kits were used: IL‐1β (Proteintech, KE10003), IL‐6 (Proteintech, KE10007), and TNF‐α (Proteintech, KE10002).

Techniques: Transplantation Assay, Irradiation, Flow Cytometry, Control, Expressing, Isolation, Derivative Assay, Marker, Two Tailed Test

Journal: Animal Cells and Systems

Article Title: Protective effect ofPhellodendri Cortexagainst lipopolysaccharide-induced memory impairment in rats

doi: 10.1080/19768354.2012.699004

Figure Lengend Snippet: Figure 5. Effects of PCE on the mean number of tumor necrosis factor-a (TNF-a)-, interleukin-1b (IL-1b)-, and cyclooxygenase- 2 (COX-2)-stained hippocampal areas after the Morris water maze test. Representative photographs and the relative percentage values are indicated in (A) and (B), respectively. (a) TNF-a expression in the CON group, (b) TNF-a expression in the LPS group, (c) TNF-a expression in the PCE200LPS group, (d) IL-1b expression in the CON group, (e) IL-1b expression in the LPS group, (f) IL-1b expression in the PCE200LPS group, (g) COX-2 expression in the CON group, (h) COX-2 expression in the LPS group, (i) COX-2 expression in the PCE200LPS group. Sections were cut coronally at 30 mm. Scale bar indicates 50 mm. **pB0.01 vs. CON group; #pB0.05 and ##pB0.01 vs. LPS group.

Article Snippet: The sections were incubated with primary rabbit anti-TNFa antibody (1:200 dilution; Novus Biochemicals LLC., Littleton, CO, USA), rabbit anti-IL-1b antibody (1:200 dilution; Santa Cruz Biotechnology Inc., Santa Cruz, CA, USA), and goat anti-COX-2 antibody (1:200 dilution; Cambridge Research Biochemicals Co., Bellingham, UK) in PBST (PBS plus 0.3% Triton X-100) for 72 h at 48C, respectively.

Techniques: Staining, Expressing

Journal: Animal Cells and Systems

Article Title: Protective effect ofPhellodendri Cortexagainst lipopolysaccharide-induced memory impairment in rats

doi: 10.1080/19768354.2012.699004

Figure Lengend Snippet: Figure 6. Effects of PCE on the expression of tumor necrosis factor-a (TNF-a), interleukin-1b (IL-1b), and cyclooxygenase-2 (COX-2) mRNAs in rats with LPS-induced hippocampal impairment. PCR bands on agarose gel and their relative intensities are indicated in (a) and (b), respectively. The expression levels of TNF-a, IL-1b, and COX-2 mRNAs were normalized to that of glyceraldehyde 3-phosphate dehydrogenase (GAPDH) mRNA as an internal control. **pB0.01 vs. CON group; #pB0.05 vs. LPS group.

Article Snippet: The sections were incubated with primary rabbit anti-TNFa antibody (1:200 dilution; Novus Biochemicals LLC., Littleton, CO, USA), rabbit anti-IL-1b antibody (1:200 dilution; Santa Cruz Biotechnology Inc., Santa Cruz, CA, USA), and goat anti-COX-2 antibody (1:200 dilution; Cambridge Research Biochemicals Co., Bellingham, UK) in PBST (PBS plus 0.3% Triton X-100) for 72 h at 48C, respectively.

Techniques: Expressing, Agarose Gel Electrophoresis, Control