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Stromal CXCL12 associates with TLS enrichment in NSCLC-BM and improved outcomes with ICI therapy. (A) Bubble plots showing chemokine receptor expression levels in Microglia, B cells, and CD4-TCF1 T cells. (B) Bubble plots CXCL12 , CXCL13 , CCL19 , and CCL21 expression levels in NSCLC-BM samples. (C) Violin plots displaying the expression levels of CXCL12 and CXCL13 in tumours from TLS-low and TLS-high groups. TLS-low group, n = 12; TLS-high group, n = 13. P < 0.001 (∗∗∗). Dashed lines indicate the median and IQR. P -value was calculated using Mann–Whitney U test. P < 0.001 (∗∗∗). (D) Representative <t>IHC</t> images of CXCL12 staining in NSCLC-BM tissues. Scale bar = 100 μm. (E) The statistical results of CXCL12 IHC staining. Upper, Correlation between stromal CXCL12 expression and TLS scores.The stromal CXCL12 IRS scores of 0–6 were classified as low expression, whereas scores of 8–12 were classified as high expression. P -value was calculated using Pearson's chi-square test. Lower, the IRS of CXCL12 expression in paired stromal regions within and outside TLSs across 52 TLS-positive samples. Data are shown as individual values with mean ± SD. P -value was calculated using Wilcoxon test. (F) Bubble plot showing CXCL12 expression levels of different cell clusters. (G) Representative mIHC images of BM tissues stained with anti-CD20 (yellow), CD31 (red), FAP (magenta), and CXCL12 (green) antibodies. Upper, scale bar = 100 μm. Lower, scale bar = 10 μm. (H) Boxplots showing the fraction of CXCL12+ cells among CD31+ cells and FAP+ cells within TLSs across 52 TLS-positive samples. Data are presented as median with IQR. P -value was calculated using the Wilcoxon test. (I) Kaplan–Meier survival analysis for iPFS in patients with NSCLC-BM receiving postoperative ICIs (n = 29) according to CXCL12 IRS in intratumoral stroma. P -value was calculated using log-rank test. NSCLC-BM, non-small cell lung cancer brain metastasis; iPFS, intracranial progression-free survival; TLS, tertiary lymphoid structure; ICIs, immune checkpoint inhibitors; IRS, immunoreactivity scores.
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Stromal CXCL12 associates with TLS enrichment in NSCLC-BM and improved outcomes with ICI therapy. (A) Bubble plots showing chemokine receptor expression levels in Microglia, B cells, and CD4-TCF1 T cells. (B) Bubble plots CXCL12 , CXCL13 , CCL19 , and CCL21 expression levels in NSCLC-BM samples. (C) Violin plots displaying the expression levels of CXCL12 and CXCL13 in tumours from TLS-low and TLS-high groups. TLS-low group, n = 12; TLS-high group, n = 13. P < 0.001 (∗∗∗). Dashed lines indicate the median and IQR. P -value was calculated using Mann–Whitney U test. P < 0.001 (∗∗∗). (D) Representative <t>IHC</t> images of CXCL12 staining in NSCLC-BM tissues. Scale bar = 100 μm. (E) The statistical results of CXCL12 IHC staining. Upper, Correlation between stromal CXCL12 expression and TLS scores.The stromal CXCL12 IRS scores of 0–6 were classified as low expression, whereas scores of 8–12 were classified as high expression. P -value was calculated using Pearson's chi-square test. Lower, the IRS of CXCL12 expression in paired stromal regions within and outside TLSs across 52 TLS-positive samples. Data are shown as individual values with mean ± SD. P -value was calculated using Wilcoxon test. (F) Bubble plot showing CXCL12 expression levels of different cell clusters. (G) Representative mIHC images of BM tissues stained with anti-CD20 (yellow), CD31 (red), FAP (magenta), and CXCL12 (green) antibodies. Upper, scale bar = 100 μm. Lower, scale bar = 10 μm. (H) Boxplots showing the fraction of CXCL12+ cells among CD31+ cells and FAP+ cells within TLSs across 52 TLS-positive samples. Data are presented as median with IQR. P -value was calculated using the Wilcoxon test. (I) Kaplan–Meier survival analysis for iPFS in patients with NSCLC-BM receiving postoperative ICIs (n = 29) according to CXCL12 IRS in intratumoral stroma. P -value was calculated using log-rank test. NSCLC-BM, non-small cell lung cancer brain metastasis; iPFS, intracranial progression-free survival; TLS, tertiary lymphoid structure; ICIs, immune checkpoint inhibitors; IRS, immunoreactivity scores.
Immunohistochemical Staining Kit, supplied by Elabscience Biotechnology, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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streptavidin biotin complex immunohistochemical staining kit  (Thermo Fisher)
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Stromal CXCL12 associates with TLS enrichment in NSCLC-BM and improved outcomes with ICI therapy. (A) Bubble plots showing chemokine receptor expression levels in Microglia, B cells, and CD4-TCF1 T cells. (B) Bubble plots CXCL12 , CXCL13 , CCL19 , and CCL21 expression levels in NSCLC-BM samples. (C) Violin plots displaying the expression levels of CXCL12 and CXCL13 in tumours from TLS-low and TLS-high groups. TLS-low group, n = 12; TLS-high group, n = 13. P < 0.001 (∗∗∗). Dashed lines indicate the median and IQR. P -value was calculated using Mann–Whitney U test. P < 0.001 (∗∗∗). (D) Representative <t>IHC</t> images of CXCL12 staining in NSCLC-BM tissues. Scale bar = 100 μm. (E) The statistical results of CXCL12 IHC staining. Upper, Correlation between stromal CXCL12 expression and TLS scores.The stromal CXCL12 IRS scores of 0–6 were classified as low expression, whereas scores of 8–12 were classified as high expression. P -value was calculated using Pearson's chi-square test. Lower, the IRS of CXCL12 expression in paired stromal regions within and outside TLSs across 52 TLS-positive samples. Data are shown as individual values with mean ± SD. P -value was calculated using Wilcoxon test. (F) Bubble plot showing CXCL12 expression levels of different cell clusters. (G) Representative mIHC images of BM tissues stained with anti-CD20 (yellow), CD31 (red), FAP (magenta), and CXCL12 (green) antibodies. Upper, scale bar = 100 μm. Lower, scale bar = 10 μm. (H) Boxplots showing the fraction of CXCL12+ cells among CD31+ cells and FAP+ cells within TLSs across 52 TLS-positive samples. Data are presented as median with IQR. P -value was calculated using the Wilcoxon test. (I) Kaplan–Meier survival analysis for iPFS in patients with NSCLC-BM receiving postoperative ICIs (n = 29) according to CXCL12 IRS in intratumoral stroma. P -value was calculated using log-rank test. NSCLC-BM, non-small cell lung cancer brain metastasis; iPFS, intracranial progression-free survival; TLS, tertiary lymphoid structure; ICIs, immune checkpoint inhibitors; IRS, immunoreactivity scores.
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signalstain ihc dual staining kit  (Cell Signaling Technology Inc)
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Stromal CXCL12 associates with TLS enrichment in NSCLC-BM and improved outcomes with ICI therapy. (A) Bubble plots showing chemokine receptor expression levels in Microglia, B cells, and CD4-TCF1 T cells. (B) Bubble plots CXCL12 , CXCL13 , CCL19 , and CCL21 expression levels in NSCLC-BM samples. (C) Violin plots displaying the expression levels of CXCL12 and CXCL13 in tumours from TLS-low and TLS-high groups. TLS-low group, n = 12; TLS-high group, n = 13. P < 0.001 (∗∗∗). Dashed lines indicate the median and IQR. P -value was calculated using Mann–Whitney U test. P < 0.001 (∗∗∗). (D) Representative <t>IHC</t> images of CXCL12 staining in NSCLC-BM tissues. Scale bar = 100 μm. (E) The statistical results of CXCL12 IHC staining. Upper, Correlation between stromal CXCL12 expression and TLS scores.The stromal CXCL12 IRS scores of 0–6 were classified as low expression, whereas scores of 8–12 were classified as high expression. P -value was calculated using Pearson's chi-square test. Lower, the IRS of CXCL12 expression in paired stromal regions within and outside TLSs across 52 TLS-positive samples. Data are shown as individual values with mean ± SD. P -value was calculated using Wilcoxon test. (F) Bubble plot showing CXCL12 expression levels of different cell clusters. (G) Representative mIHC images of BM tissues stained with anti-CD20 (yellow), CD31 (red), FAP (magenta), and CXCL12 (green) antibodies. Upper, scale bar = 100 μm. Lower, scale bar = 10 μm. (H) Boxplots showing the fraction of CXCL12+ cells among CD31+ cells and FAP+ cells within TLSs across 52 TLS-positive samples. Data are presented as median with IQR. P -value was calculated using the Wilcoxon test. (I) Kaplan–Meier survival analysis for iPFS in patients with NSCLC-BM receiving postoperative ICIs (n = 29) according to CXCL12 IRS in intratumoral stroma. P -value was calculated using log-rank test. NSCLC-BM, non-small cell lung cancer brain metastasis; iPFS, intracranial progression-free survival; TLS, tertiary lymphoid structure; ICIs, immune checkpoint inhibitors; IRS, immunoreactivity scores.
Signalstain Ihc Dual Staining Kit, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Stromal CXCL12 associates with TLS enrichment in NSCLC-BM and improved outcomes with ICI therapy. (A) Bubble plots showing chemokine receptor expression levels in Microglia, B cells, and CD4-TCF1 T cells. (B) Bubble plots CXCL12 , CXCL13 , CCL19 , and CCL21 expression levels in NSCLC-BM samples. (C) Violin plots displaying the expression levels of CXCL12 and CXCL13 in tumours from TLS-low and TLS-high groups. TLS-low group, n = 12; TLS-high group, n = 13. P < 0.001 (∗∗∗). Dashed lines indicate the median and IQR. P -value was calculated using Mann–Whitney U test. P < 0.001 (∗∗∗). (D) Representative IHC images of CXCL12 staining in NSCLC-BM tissues. Scale bar = 100 μm. (E) The statistical results of CXCL12 IHC staining. Upper, Correlation between stromal CXCL12 expression and TLS scores.The stromal CXCL12 IRS scores of 0–6 were classified as low expression, whereas scores of 8–12 were classified as high expression. P -value was calculated using Pearson's chi-square test. Lower, the IRS of CXCL12 expression in paired stromal regions within and outside TLSs across 52 TLS-positive samples. Data are shown as individual values with mean ± SD. P -value was calculated using Wilcoxon test. (F) Bubble plot showing CXCL12 expression levels of different cell clusters. (G) Representative mIHC images of BM tissues stained with anti-CD20 (yellow), CD31 (red), FAP (magenta), and CXCL12 (green) antibodies. Upper, scale bar = 100 μm. Lower, scale bar = 10 μm. (H) Boxplots showing the fraction of CXCL12+ cells among CD31+ cells and FAP+ cells within TLSs across 52 TLS-positive samples. Data are presented as median with IQR. P -value was calculated using the Wilcoxon test. (I) Kaplan–Meier survival analysis for iPFS in patients with NSCLC-BM receiving postoperative ICIs (n = 29) according to CXCL12 IRS in intratumoral stroma. P -value was calculated using log-rank test. NSCLC-BM, non-small cell lung cancer brain metastasis; iPFS, intracranial progression-free survival; TLS, tertiary lymphoid structure; ICIs, immune checkpoint inhibitors; IRS, immunoreactivity scores.

Journal: eBioMedicine

Article Title: TCF1+CD4+ T cells and microglia co-orchestrate tertiary lymphoid structures to enhance prognosis and immunotherapy in non-small cell lung cancer with brain metastases

doi: 10.1016/j.ebiom.2026.106280

Figure Lengend Snippet: Stromal CXCL12 associates with TLS enrichment in NSCLC-BM and improved outcomes with ICI therapy. (A) Bubble plots showing chemokine receptor expression levels in Microglia, B cells, and CD4-TCF1 T cells. (B) Bubble plots CXCL12 , CXCL13 , CCL19 , and CCL21 expression levels in NSCLC-BM samples. (C) Violin plots displaying the expression levels of CXCL12 and CXCL13 in tumours from TLS-low and TLS-high groups. TLS-low group, n = 12; TLS-high group, n = 13. P < 0.001 (∗∗∗). Dashed lines indicate the median and IQR. P -value was calculated using Mann–Whitney U test. P < 0.001 (∗∗∗). (D) Representative IHC images of CXCL12 staining in NSCLC-BM tissues. Scale bar = 100 μm. (E) The statistical results of CXCL12 IHC staining. Upper, Correlation between stromal CXCL12 expression and TLS scores.The stromal CXCL12 IRS scores of 0–6 were classified as low expression, whereas scores of 8–12 were classified as high expression. P -value was calculated using Pearson's chi-square test. Lower, the IRS of CXCL12 expression in paired stromal regions within and outside TLSs across 52 TLS-positive samples. Data are shown as individual values with mean ± SD. P -value was calculated using Wilcoxon test. (F) Bubble plot showing CXCL12 expression levels of different cell clusters. (G) Representative mIHC images of BM tissues stained with anti-CD20 (yellow), CD31 (red), FAP (magenta), and CXCL12 (green) antibodies. Upper, scale bar = 100 μm. Lower, scale bar = 10 μm. (H) Boxplots showing the fraction of CXCL12+ cells among CD31+ cells and FAP+ cells within TLSs across 52 TLS-positive samples. Data are presented as median with IQR. P -value was calculated using the Wilcoxon test. (I) Kaplan–Meier survival analysis for iPFS in patients with NSCLC-BM receiving postoperative ICIs (n = 29) according to CXCL12 IRS in intratumoral stroma. P -value was calculated using log-rank test. NSCLC-BM, non-small cell lung cancer brain metastasis; iPFS, intracranial progression-free survival; TLS, tertiary lymphoid structure; ICIs, immune checkpoint inhibitors; IRS, immunoreactivity scores.

Article Snippet: IHC staining was performed using the SP method, employing the universal immunohistochemical staining kit (Zhongshan Golden Bridge Biotechnology; Cat# PV-6000) and DAB chromogen kit (Zhongshan Golden Bridge Biotechnology; Cat# PV-6000D).

Techniques: Expressing, MANN-WHITNEY, Staining, Immunohistochemistry