Journal: JVS-Vascular Science

Article Title: EphB4 monomer inhibits chronic graft vasculopathy in an aortic transplant model

doi: 10.1016/j.jvssci.2023.100109

Figure Lengend Snippet: EphB4 monomer reduces CD68 + cell infiltration in allografts. (A) Representative immunofluorescence of isografts and allografts (day 28) treated with either control IgG or EphB4 monomer; top row, 40× (scale bar, 250 μm); bottom row, 400× (scale bar, 25 μm). (B) Mean number of CD68 + cells per high-power field ( HPF ) in isografts; P = .66 (Mann-Whitney U test; n = 5-6 rats per group). (C) Mean number of CD68 + cells per HPF in allografts; P ≤ .01 (Mann-Whitney U test; n = 5-6 rats per group).

Article Snippet: Rats then had subdermal injections with either EphB4 monomer (20 μg/kg; Sino Biological Inc., Beijing, China) or IgG control (Sino Biological Inc.).

Techniques: Immunofluorescence, MANN-WHITNEY

Journal: JVS-Vascular Science

Article Title: EphB4 monomer inhibits chronic graft vasculopathy in an aortic transplant model

doi: 10.1016/j.jvssci.2023.100109

Figure Lengend Snippet: EphB4 monomer reduces CD3 + cell infiltration in allografts. (A) Representative immunofluorescence of isografts and allografts (day 28) treated with either control IgG or EphB4 monomer; top row, 40× (scale bar, 250 μm); bottom row, 400× (scale bar, 25 μm). (B) Mean number of CD3 + cells per high-power field ( HPF ) in isografts ( P = .17, Mann-Whitney U test ; n = 5-6 rats per group). (C) Mean number of CD3 + cells per HPF in allografts; P = .02 (Mann-Whitney U test; n = 5-6 rats per group).

Article Snippet: Rats then had subdermal injections with either EphB4 monomer (20 μg/kg; Sino Biological Inc., Beijing, China) or IgG control (Sino Biological Inc.).

Techniques: Immunofluorescence, MANN-WHITNEY

Journal: JVS-Vascular Science

Article Title: EphB4 monomer inhibits chronic graft vasculopathy in an aortic transplant model

doi: 10.1016/j.jvssci.2023.100109

Figure Lengend Snippet: EphB4 monomer reduces neointimal thickness in allografts. (A) Representative EVG staining of isografts and allografts (day 28) treated with either control IgG or EphB4 monomer; top row, 40× (scale bar, 250 μm); bottom row, 400× (scale, bar 25 μm). (B) Mean neointimal thickness in isografts ( P = .08, Mann-Whitney U test; n = 5-6 rats per group). (C) Mean medial thickness in isografts ( P = .08, Mann-Whitney U test; n = 5-6 rats per group). (D) Ratio of neointima:media thickness in isografts ( P = .66, Mann-Whitney U test; n = 5-6 rats per group). (E) Mean neointimal thickness in allografts ( P = .05, Mann-Whitney U test; n = 5-6 rats per group). (F) Mean medial thickness in allografts ( P = .12, Mann-Whitney U test; n = 5-6 rats per group). (G) Ratio of neointima:media thickness in allografts ( P = .05, Mann-Whitney U test; n = 5-6 rats per group).

Article Snippet: Rats then had subdermal injections with either EphB4 monomer (20 μg/kg; Sino Biological Inc., Beijing, China) or IgG control (Sino Biological Inc.).

Techniques: Staining, MANN-WHITNEY

Journal: Clinical and Translational Medicine

Article Title: Single‐cell transcriptome sequencing of B‐cell heterogeneity and tertiary lymphoid structure predicts breast cancer prognosis and neoadjuvant therapy efficacy

doi: 10.1002/ctm2.1346

Figure Lengend Snippet: Correlation analysis between TIL‐B and mature tertiary lymphoid structures (TLS). (A) Among the 200 patients included in the BRCA dataset from the TCGA research consortium, a subset of 18 patients who exhibited TLS positivity were identified. H&E images of characteristic TLS in these patients. (B) Co‐immunofluorescence subimages depicting the co‐localization of CD23 and CD21 within CD23‐positive TLS were obtained from a collection of 70 BC samples that we compiled. (C) In 9 TLS‐positive patients of 70 BC patients, representative images of H&E staining and IHC staining of CD4, CD8, CD20, CD23 and BCL6 expression in TLS‐positive BC patients. (D) Analysis of DEGs in 200 TCGA BRCA cases categorized as TLS‐negative and TLS‐positive. Source data are available at: https://portal.gdc.cancer.gov . (E and F) H&E (E) and CD23 IHC (F) images of the typical TLS‐positive case in 70 BC patients. (G) Different tumour types exhibit FCER2 expression, according to TIMER2. * p < .05;** p < .01;*** p < .001. (H) Utilizing the EPIC and TIDE algorithms within TIMER2, the relationship between FCER2 expression and tumour infiltrating immune cells in BC was computed. (I‐J) Kaplan–Meier Survival curve of FCER2(I), co‐acting with FCER2 and B cells(J) in BC by TIMER2. TIL‐B, tumor‐infiltrating B cells.

Article Snippet: 1% bovine serum albumin for 10 min at room temperature, followed by overnight incubation at 4°C with the following primary antibodies: monoclonal antibody against FCER2 (CD23; 1:400; 60208‐2‐Ig; Proteintech), anti‐CD21 (1:400; 24374‐1‐AP; Proteintech), anti‐CD20 (1:400; 60271‐1‐Ig; Proteintech), anti‐CD4 (1:400; 67786‐1‐Ig; Proteintech), anti‐CD8 (1:400; 66868‐1‐Ig; Proteintech) and anti‐BCL6 (1:400; 66340‐1‐Ig; Proteintech).

Techniques: Immunofluorescence, Staining, Immunohistochemistry, Expressing

Journal: Clinical and Translational Medicine

Article Title: Single‐cell transcriptome sequencing of B‐cell heterogeneity and tertiary lymphoid structure predicts breast cancer prognosis and neoadjuvant therapy efficacy

doi: 10.1002/ctm2.1346

Figure Lengend Snippet: Different tertiary lymphoid structures (TLS) states in breast cancer (BC) display metabolic variability and survival variations. (A) A bar graph depicting the distribution of immune cells that have infiltrated tumours in TLS‐positive and TLS‐negative groups among 9 BC patients. (B) Genetic analysis of immune cell differences in TLS‐positive and TLS‐negative groups among 9 BC patients. (C) Heatmap of metabolic pathway scores in TLS‐positive and TLS‐negative groups among 9 BC patients. (D) Heatmap showing average metabolic gene expression in TLS‐positive and TLS‐negative groups among 9 BC patients. (E) To ascertain the expression correlation between CD23 and the biomarkers citrate synthase (CS), HK2, lactate dehydrogenase A (LDHA), and IDH3G in BC, the cBioportal tool ( https://www.cbioportal.org ) was utilized, employing the TCGA dataset (Firehose Legacy dataset; encompassing 1108 BC samples) for this purpose. (F) Kaplan–Meier survival curves depicting the disparity in BC outcomes between cohorts with high and low APOD expression, as analyzed through GEPIA. (G) Imaging before and after neoadjuvant chemotherapy of TLS‐positive patients with significant effectiveness. (H) Imaging before and after neoadjuvant immunotherapy of TLS‐positive patients with significant effectiveness.

Article Snippet: 1% bovine serum albumin for 10 min at room temperature, followed by overnight incubation at 4°C with the following primary antibodies: monoclonal antibody against FCER2 (CD23; 1:400; 60208‐2‐Ig; Proteintech), anti‐CD21 (1:400; 24374‐1‐AP; Proteintech), anti‐CD20 (1:400; 60271‐1‐Ig; Proteintech), anti‐CD4 (1:400; 67786‐1‐Ig; Proteintech), anti‐CD8 (1:400; 66868‐1‐Ig; Proteintech) and anti‐BCL6 (1:400; 66340‐1‐Ig; Proteintech).

Techniques: Gene Expression, Expressing, Imaging

Journal: Clinical and Translational Medicine

Article Title: Single‐cell transcriptome sequencing of B‐cell heterogeneity and tertiary lymphoid structure predicts breast cancer prognosis and neoadjuvant therapy efficacy

doi: 10.1002/ctm2.1346

Figure Lengend Snippet: The potential significance of tertiary lymphoid structures (TLS) and TLS‐specific markers for breast cancer (BC) prognosis. After matching with Fitness score matching (PSM), survival analysis and prognostic factor analysis were performed. For survival analysis, the Kaplan–Meier method and the log‐rank test were employed. For prognostic factors, COX proportional hazard regression model was used. The two‐sided log rank test was used to determine the p values. (A) Kaplan–Meier survival curves for the disease‐free survival (DFS) (left) and OS (right) of 920 TCGA BRCA patients based on single gene expression (CD20, CD23 and CD8). (B) The impact of CD20, CD23, CD8, CD4 and BCL6 on 920 TCGA BRCA patients’ prognosis. The forest map shows HRs (center pink and blue squares) and 95% confidence interval (horizontal ranges), and PSM matching has been made for factors such as molecular typing, lymph node status, tumour size, diagnosis age and histological grading of BC. (C) Kaplan–Meier survival curves for the DFS of 70 BC patients based on TLS expression.

Article Snippet: 1% bovine serum albumin for 10 min at room temperature, followed by overnight incubation at 4°C with the following primary antibodies: monoclonal antibody against FCER2 (CD23; 1:400; 60208‐2‐Ig; Proteintech), anti‐CD21 (1:400; 24374‐1‐AP; Proteintech), anti‐CD20 (1:400; 60271‐1‐Ig; Proteintech), anti‐CD4 (1:400; 67786‐1‐Ig; Proteintech), anti‐CD8 (1:400; 66868‐1‐Ig; Proteintech) and anti‐BCL6 (1:400; 66340‐1‐Ig; Proteintech).

Techniques: Gene Expression, Biomarker Discovery, Expressing

Journal: Alzheimer's & dementia : the journal of the Alzheimer's Association

Article Title: Peripheral complement interactions with amyloid β peptide (Aβ) in Alzheimer’s disease: polymorphisms, structure, and function of complement receptor 1

doi: 10.1016/j.jalz.2018.04.003

Figure Lengend Snippet: A) Direct binding and degradation of pathogens such as Aβ can occur without CR1 mediation, but is typically less effective than CR1-dependent mechanisms [52]. B) Classical immune adherence occurs when pathogen/antibody immune complexes activate complement, resulting in fixation of the immune complex with complement opsonins, particularly C3b. C3b is a primary ligand for CR1 and therefore binds the immune complex to CR1 on monocyte/macrophage and erythrocyte surfaces. Such binding is considered to be more effective/facile than direct binding [52], as is the case for Aβ [26]. After capture of the pathogen/immune complex, monocyte/macrophages then internalize and degrade the pathogen. Erythrocytes, however, ferry the bound immune complex to the liver, where the pathogen is stripped off by Kupffer cells and degraded [22,23]. C) Aβ in its aggregated state is one of a handful of peptides that can activate complement without antibody mediation [24–30]. As a result, it is tagged with complement opsonins such as C3b [29,30] and becomes subject to capture by CR1. D) The most efficacious binding and degradation of pathogens occurs when both antibody-dependent (Panel B) and antibody-independent (Panel C) mechanisms of complement opsonization occur (Panel D) [52].

Article Snippet: C___8828478_10 , CR2;CR1 , 1 , rs1571344.

Techniques: Binding Assay

Journal: Alzheimer's & dementia : the journal of the Alzheimer's Association

Article Title: Peripheral complement interactions with amyloid β peptide (Aβ) in Alzheimer’s disease: polymorphisms, structure, and function of complement receptor 1

doi: 10.1016/j.jalz.2018.04.003

Figure Lengend Snippet: CR1 is synthesized in four different structural isoforms. The F (fast migrating on electrophoresis) isoform has a molecular weight of ~190 kDa; the S (slow migrating on electrophoresis) isoform has a molecular weight of ~220 kDa. The C (~160 kDa) and D (~250 kDa) isoforms are quite rare (<1% of the population) [43]. Western blots with CR1 clone E11 antibody reveal these structural isoforms. For example, the patient in lane 3 (F/C) has one allele that expresses the F isoform and one allele that expresses the C isoform, whereas the patients in lanes 4–6 (F/F) are homozygous for the F isoform, and the patient in lane 9 (S/S) is homozygous for the S isoform. In addition to defining the structural isoform genotype for each patient, Western blots of CR1 have the further advantage of providing separate expression measures for each isoform (i.e., densitometry of each isoform b and) as well as measures of total erythrocyte CR1 (i.e., sum of the levels for each band). (MW) Molecular weight standards. (rCR1) Recombinant CR1.

Article Snippet: C___8828478_10 , CR2;CR1 , 1 , rs1571344.

Techniques: Synthesized, Electrophoresis, Molecular Weight, Western Blot, Expressing, Recombinant

Journal: Alzheimer's & dementia : the journal of the Alzheimer's Association

Article Title: Peripheral complement interactions with amyloid β peptide (Aβ) in Alzheimer’s disease: polymorphisms, structure, and function of complement receptor 1

doi: 10.1016/j.jalz.2018.04.003

Figure Lengend Snippet: CR1 and A po E SNPs evaluated in the present study for their relationships to erythrocyte clearance mechanisms and AD risk

Article Snippet: C___8828478_10 , CR2;CR1 , 1 , rs1571344.

Techniques:

Journal: Alzheimer's & dementia : the journal of the Alzheimer's Association

Article Title: Peripheral complement interactions with amyloid β peptide (Aβ) in Alzheimer’s disease: polymorphisms, structure, and function of complement receptor 1

doi: 10.1016/j.jalz.2018.04.003

Figure Lengend Snippet: A) Representative immunocytochemistry of erythrocyte ghosts from a normal elderly subject using a conventional, commercial CR1 antibody (#376223, clone E11, US Biologicals, Salem, MA) directed at an epitope in the C3b/C4b binding region of CR1. CR1 immunoreactivity was readily detected at the cell surface and in clusters, characteristics of this cell surface receptor [53]. B) Representative immunohistochemistry of a <4 hours postmortem cortical brain section from an AD case using the same antibody and conditions as in Panel A. No material staining of any cell type was observed in any of the 4 AD or 4 ND patients studied, except for staining of cells that clearly lay in the brain vasculature (inset). Similar results were obtained with the three other CR1 antibodies employed in the experiments. C) In addition to erythrocytes, other blood cells known to express CR1 were detected in the brain vasculature by these methods. For example, CR1 immunoreactive eosinophils could be identified based on their multi-nucleated morphology (inset). D) In Western blots with CR1 clone E11 antibody and conventional 20 μg/ml/lane protein loads, CR1 immunoreactive bands were readily detected in erythrocyte samples (E) (lane 10) at the same molecular weight as recombinant CR1 (rCR1) (lane 9). Under the same conditions, only faint traces of immunoreactive CR1 were observed for the neocortical samples even at extremely high, 100 μg/ml/lane loads. E) Nearly identical results were obtained for the same samples in parallel blots of GPVI, a platelet marker that is expressly not found in brain [54], strongly suggesting that it (and putative brain CR1) are simply vascular contaminants that are inherently trapped in brain homogenates. Autofluorescent and immunoreactive hemoglobin could also be observed at these high protein concentrations (not shown). (P) Plasma sample showing bands for GPVI, which appear as multimers of the 62 kDa monomer form [55].

Article Snippet: C___8828478_10 , CR2;CR1 , 1 , rs1571344.

Techniques: Immunocytochemistry, Binding Assay, Cell Surface Receptor Assay, Immunohistochemistry, Staining, Western Blot, Molecular Weight, Recombinant, Marker, Clinical Proteomics

Journal: Alzheimer's & dementia : the journal of the Alzheimer's Association

Article Title: Peripheral complement interactions with amyloid β peptide (Aβ) in Alzheimer’s disease: polymorphisms, structure, and function of complement receptor 1

doi: 10.1016/j.jalz.2018.04.003

Figure Lengend Snippet: A) Log plot of qPCR for CR1 mRNA and mRNAs for other blood and brain proteins in brain samples. At 30 amplification cycles, mRNAs for brain-specific markers such as GFAP and ALDH1L1 were readily detected in brain homogenates from four AD and four ND subjects, whereas, in the same samples, there was no detectable mRNA for CR1 with either of two different primers. Likewise, mRNAs for the hemoglobin marker HBA2 [42] exhibited only trace values. Nonetheless, in these same samples, small but consistent amounts of mRNAs for CR1 and HBA2 began to be observed after 40 amplification cycles, although their copy numbers were still some 2 orders of magnitude less than those for the brain-specific markers. B) As expected, using the same primers, mRNAs for CR1 and HBA2 were readily detected in blood samples at both 30 and 40 amplification cycles. By contrast, neither of the brain-specific markers had detectable mRNAs at 30 amplification cycles, and only trace amounts of astrocyte-specific GFAP mRNA were observed at 40 cycles. C) In single cell laser captured microglia (N = 21,600) from 6 AD, 6 ND, and 6 PD cases, CR1 mRNA expression was essentially background (mean FPKM = 0.04 ± 0.03), with 0 values for 94% of samples, whereas mRNAs for other markers known to be expressed by microglia were readily observed in all samples.

Article Snippet: C___8828478_10 , CR2;CR1 , 1 , rs1571344.

Techniques: Amplification, Marker, Expressing

Journal: Alzheimer's & dementia : the journal of the Alzheimer's Association

Article Title: Peripheral complement interactions with amyloid β peptide (Aβ) in Alzheimer’s disease: polymorphisms, structure, and function of complement receptor 1

doi: 10.1016/j.jalz.2018.04.003

Figure Lengend Snippet: A) Erythrocyte CR1 expression as measured by densitometry of Western blots using AD and ND erythrocyte membrane preparations. B) Erythrocyte Aβ levels as measured by flow cytometry. These data confirm our earlier reports of deficient erythrocyte Aβ capture using ELISA methods [24–26]. C) Relationship of erythrocyte CR1 expression to the structural isoforms of CR1. Possession of the F isoform in one or both alleles significantly increased total erythrocyte CR1 levels, in keeping with previous findings [16]. Homozygosity for the F allele (F/F) was particularly potent in this regard. D) Homozygosity for the CR1-enhancing F/F genotype was significantly under-represented in AD compared to ND subjects. E) Likewise, expression levels of the F isoform overall were significantly deficient in AD compared to ND subjects.

Article Snippet: C___8828478_10 , CR2;CR1 , 1 , rs1571344.

Techniques: Expressing, Western Blot, Membrane, Flow Cytometry, Enzyme-linked Immunosorbent Assay