856163f035f892efccf518197b859ae2 pmc_34071393.pdf 054e4ef4a543bd120cc347027e3c9b6df11346d7 pmc_34071393.pdf e5530aa268266eea32a97fddaeb4648129df23e492c7b30a8f67293c1ce95c37 pmc_34071393.pdf Title: Immune System-Related Changes in Preclinical GL261 Glioblastoma under TMZ Treatment: Explaining MRSI-Based Nosological Imaging Findings with RT-PCR Analyses Subject: Glioblastomas (GB) are brain tumours with poor prognosis even after aggressive therapy. Previous work suggests that magnetic resonance spectroscopic imaging (MRSI) could act as a biomarker of efficient immune system attack onto GB, presenting oscillatory changes. Glioma-associated microglia/macrophages (GAMs) constitute the most abundant non-tumour cell type within the GB and can be polarised into anti-tumour (M1) or pro-tumour (M2) phenotypes. One of the mechanisms to mediate immunosuppression in brain tumours is the interaction between programmed cell death-1 ligand 1 (PD-L1) and programmed cell death-1 receptor (PD-1). We evaluated the subpopulations of GAMs in responding and control GB tumours to correlate PD-L1 expression to GAM polarisation in order to explain/validate MRSI-detected findings. Mice were evaluated by MRI/MRSI to assess the extent of response to treatment and with qPCR for GAMs M1 and M2 polarisation analyses. M1/M2 ratios and PD-L1 expression were higher in treated compared to control tumours. Furthermore, PD-L1 expression was positively correlated with the M1/M2 ratio. The oscillatory change in the GAMs prevailing population could be one of the key causes for the differential MRSI-detected pattern, allowing this to act as immune system activity biomarker in future work. Keywords: glioblastoma; orthotopic immunocompetent tumours; immune-enhancing metronomic schedule; TMZ; magnetic resonance spectroscopic imaging; immune system activity imaging biomarker; cancer immune cycle; glioma-associated microglia/macrophages; PD-L1 Author: Pilar Calero-Pérez, Shuang Wu, Carles Arús and Ana Paula Candiota Creator: LaTeX with hyperref Producer: pdfTeX-1.40.21 CreationDate: Mon Jun 7 02:47:18 2021 CEST ModDate: Mon Jun 7 04:48:17 2021 CEST Custom Metadata: no Metadata Stream: no Tagged: no UserProperties: no Suspects: no Form: none JavaScript: no Pages: 19 Encrypted: no Page size: 595.276 x 841.89 pts (A4) Page rot: 0 File size: 1749525 bytes Optimized: no PDF version: 1.7 name type encoding emb sub uni object ID ------------------------------------ ----------------- ---------------- --- --- --- --------- NQWDWO+VnURWPalladioL Type 1 Custom yes yes no 10 0 CDXJMD+URWPalladioL-Ital Type 1 Custom yes yes no 15 0 QIHDIK+URWPalladioL-Bold Type 1 Custom yes yes no 20 0 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DMBMOF+PalatinoLinotype TrueType WinAnsi yes yes no 153 0 DMBMOH+PalatinoLinotype,Italic TrueType WinAnsi yes yes no 156 0 DMBMOD+PalatinoLinotype,Bold TrueType WinAnsi yes yes no 170 0 DMBMOF+PalatinoLinotype TrueType WinAnsi yes yes no 173 0 DMBMOH+PalatinoLinotype,Italic TrueType WinAnsi yes yes no 176 0 DMBMOD+PalatinoLinotype,Bold TrueType WinAnsi yes yes no 190 0 DMBMOF+PalatinoLinotype TrueType WinAnsi yes yes no 193 0 DMBMOH+PalatinoLinotype,Italic TrueType WinAnsi yes yes no 196 0 Jhove (Rel. 1.28.0, 2023-05-18) Date: 2024-10-02 03:15:15 CEST RepresentationInformation: pmc_34071393.pdf ReportingModule: PDF-hul, Rel. 1.12.4 (2023-03-16) LastModified: 2023-03-31 12:14:04 CEST Size: 1749525 Format: PDF Version: 1.7 Status: Well-Formed and valid SignatureMatches: PDF-hul MIMEtype: application/pdf PDFMetadata: Objects: 565 FreeObjects: 1 IncrementalUpdates: 0 DocumentCatalog: PageLayout: SinglePage PageMode: UseNone Outlines: Item: Title: Introduction Destination: section.1 Item: Title: Materials and Methods Destination: section.2 Children: Item: Title: GL261 GB Preclinical Model Generation and Treatment Destination: subsection.2.1 Item: Title: In Vivo MRI and MRSI Studies Destination: subsection.2.2 Children: Item: Title: MRI Studies Destination: subsubsection.2.2.1 Item: Title: MRSI Studies Destination: subsubsection.2.2.2 Item: Title: MRI and MRSI Processing and Post-Processing Destination: subsubsection.2.2.3 Item: Title: Tumour Responding index (TRI) Calculations Destination: subsubsection.2.2.4 Item: Title: Animal Euthanasia Destination: subsection.2.3 Item: Title: RNA Isolation, cDNA Synthesis and qPCR Destination: subsection.2.4 Item: Title: Statistical Analysis Destination: subsection.2.5 Item: Title: Results Destination: section.3 Children: Item: Title: Follow up of GL261 Tumour-Bearing Mice and Endpoint Criteria Destination: subsection.3.1 Item: Title: Microglia/Macrophage Global Population, As Well As M1 and M2 Subtypes, Are Increased in IMS-TMZ-Treated Tumours Destination: subsection.3.2 Item: Title: Assessing Different Macrophage Population Subtypes Regarding Global GAMs Values Destination: subsection.3.3 Item: Title: PD-L1 Gene Is Highly Expressed in IMS-TMZ-Treated Tumours, and These Increases May Be Correlated with the Polarisation State of Microglia/Macrophage Population Destination: subsection.3.4 Item: Title: Discussion Destination: section.4 Children: Item: Title: Immune System Populations Change during Response to Therapy Destination: subsection.4.1 Item: Title: Not All GAMs Are Equal (I): Polarisation of Microglia/Macrophages and Its Role in Therapy Response in GB Destination: subsection.4.2 Item: Title: Not All GAMs Are Equal (II): The Polarisation Status of Microglia/Macrophages Affects Their Metabolomic Pattern Destination: subsection.4.3 Item: Title: PD-L1 Gene Expression in GL261 GB Destination: subsection.4.4 Item: Title: Wrap-up: Incoporating the Measured Gene Expression Results into the Explanation of the Oscillatory Behaviour of the MRSI-Based Biomarker of Response Destination: subsection.4.5 Item: Title: Conclusions Destination: section.5 Item: Title: References Destination: section.5 Info: Title: Immune System-Related Changes in Preclinical GL261 Glioblastoma under TMZ Treatment: Explaining MRSI-Based Nosological Imaging Findings with RT-PCR Analyses Author: Pilar Calero-Pérez, Shuang Wu, Carles Arús and Ana Paula Candiota Subject: Glioblastomas (GB) are brain tumours with poor prognosis even after aggressive therapy. Previous work suggests that magnetic resonance spectroscopic imaging (MRSI) could act as a biomarker of efficient immune system attack onto GB, presenting oscillatory changes. Glioma-associated microglia/macrophages (GAMs) constitute the most abundant non-tumour cell type within the GB and can be polarised into anti-tumour (M1) or pro-tumour (M2) phenotypes. One of the mechanisms to mediate immunosuppression in brain tumours is the interaction between programmed cell death-1 ligand 1 (PD-L1) and programmed cell death-1 receptor (PD-1). We evaluated the subpopulations of GAMs in responding and control GB tumours to correlate PD-L1 expression to GAM polarisation in order to explain/validate MRSI-detected findings. Mice were evaluated by MRI/MRSI to assess the extent of response to treatment and with qPCR for GAMs M1 and M2 polarisation analyses. M1/M2 ratios and PD-L1 expression were higher in treated compared to control tumours. Furthermore, PD-L1 expression was positively correlated with the M1/M2 ratio. The oscillatory change in the GAMs prevailing population could be one of the key causes for the differential MRSI-detected pattern, allowing this to act as immune system activity biomarker in future work. Keywords: glioblastoma; orthotopic immunocompetent tumours; immune-enhancing metronomic schedule; TMZ; magnetic resonance spectroscopic imaging; immune system activity imaging biomarker; cancer immune cycle; glioma-associated microglia/macrophages; PD-L1 Creator: LaTeX with hyperref Producer: pdfTeX-1.40.21 CreationDate: Mon Jun 07 02:47:18 CEST 2021 ModDate: Mon Jun 07 04:48:17 CEST 2021 ID: 0x916f5d754454df66d646e0c64ab1f5b1, 0x916f5d754454df66d646e0c64ab1f5b1 Filters: FilterPipeline: FlateDecode Fonts: Type1: Font: BaseFont: JHYTSG+CMR10 FontSubset: true FirstChar: 61 LastChar: 61 FontDescriptor: FontName: JHYTSG+CMR10 Flags: Symbolic FontBBox: -40, -250, 1009, 750 FontFile: true Font: BaseFont: IDSVYB+VnURWPalladioL-Italic FontSubset: true FirstChar: 145 LastChar: 226 FontDescriptor: FontName: IDSVYB+VnURWPalladioL-Italic Flags: Symbolic FontBBox: -170, -305, 1010, 941 FontFile: true EncodingDictionary: Differences: true Font: BaseFont: QIHDIK+URWPalladioL-Bold FontSubset: true 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