Tumor-associated macrophages (TAMs) plentifully infiltrate the tumor microenvironment (TME), but their role in anti-tumor immunity is controversial. Depending on the acquired polarization, they can either support tumor growth or participate in the elimination of neoplastic cells. In this study, we analyzed the TME by RNA-seq and flow cytometry and examined TAMs after ex vivo activation. Tumors with normal and either reversibly or irreversibly decreased expression of major histocompatibility complex class I (MHC-I) molecules were induced with TC-1, TC-1/A9, and TC-1/dB2m cells, respectively. We found that combined immunotherapy (IT), composed of DNA immunization and the CpG oligodeoxynucleotide (ODN) ODN1826, evoked immune reactions in the TME of TC-1- and TC-1/A9-induced tumors, while the TME of TC-1/dB2m tumors was mostly immunologically unresponsive. TAMs infiltrated both tumor types with MHC-I downregulation, but only TAMs from TC-1/A9 tumors acquired the M1 phenotype upon IT and were cytotoxic in in vitro assay. The anti-tumor effect of combined IT was markedly enhanced by a blockade of the colony-stimulating factor-1 receptor (CSF-1R), but only against TC-1/A9 tumors. Overall, TAMs from tumors with irreversible MHC-I downregulation were resistant to the stimulation of cytotoxic activity. These data suggest the dissimilarity of TAMs from different tumor types, which should be considered when utilizing TAMs in cancer IT.

• Publikační typ
• časopisecké články MeSH

BACKGROUND: Vertebrate meiotic recombination events are concentrated in regions (hotspots) that display open chromatin marks, such as trimethylation of lysines 4 and 36 of histone 3 (H3K4me3 and H3K36me3). Mouse and human PRDM9 proteins catalyze H3K4me3 and H3K36me3 and determine hotspot positions, whereas other vertebrates lacking PRDM9 recombine in regions with chromatin already opened for another function, such as gene promoters. While these other vertebrate species lacking PRDM9 remain fertile, inactivation of the mouse Prdm9 gene, which shifts the hotspots to the functional regions (including promoters), typically causes gross fertility reduction; and the reasons for these species differences are not clear. RESULTS: We introduced Prdm9 deletions into the Rattus norvegicus genome and generated the first rat genome-wide maps of recombination-initiating double-strand break hotspots. Rat strains carrying the same wild-type Prdm9 allele shared 88% hotspots but strains with different Prdm9 alleles only 3%. After Prdm9 deletion, rat hotspots relocated to functional regions, about 40% to positions corresponding to Prdm9-independent mouse hotspots, including promoters. Despite the hotspot relocation and decreased fertility, Prdm9-deficient rats of the SHR/OlaIpcv strain produced healthy offspring. The percentage of normal pachytene spermatocytes in SHR-Prdm9 mutants was almost double than in the PWD male mouse oligospermic sterile mutants. We previously found a correlation between the crossover rate and sperm presence in mouse Prdm9 mutants. The crossover rate of SHR is more similar to sperm-carrying mutant mice, but it did not fully explain the fertility of the SHR mutants. Besides mild meiotic arrests at rat tubular stages IV (mid-pachytene) and XIV (metaphase), we also detected postmeiotic apoptosis of round spermatids. We found delayed meiosis and age-dependent fertility in both sexes of the SHR mutants. CONCLUSIONS: We hypothesize that the relative increased fertility of rat versus mouse Prdm9 mutants could be ascribed to extended duration of meiotic prophase I. While rat PRDM9 shapes meiotic recombination landscapes, it is unnecessary for recombination. We suggest that PRDM9 has additional roles in spermatogenesis and speciation-spermatid development and reproductive age-that may help to explain male-specific hybrid sterility.

• MeSH
• chromatin MeSH
• dvouřetězcové zlomy DNA MeSH
• fertilita genetika   MeSH
• histonlysin-N-methyltransferasa genetika   MeSH
• krysa rodu rattus MeSH
• meióza * genetika   MeSH
• myši MeSH
• potkani inbrední SHR MeSH
• spermatogeneze genetika   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• mužské pohlaví MeSH
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH

Cluster of differentiation (CD) 80 is mainly expressed in immune cells but can also be found in several types of cancer cells. This molecule may either activate or inhibit immune reactions. Here, we determined the immunosuppressive role of CD80 in the tumor microenvironment by CRISPR/Cas9-mediated deactivation of the corresponding gene in the mouse oncogenic TC-1 cell line. The tumor cells with deactivated CD80 (TC-1/dCD80-1) were more immunogenic than parental cells and induced tumors that gained sensitivity to cytotoxic T-lymphocyte antigen 4 (CTLA-4) blockade, as compared with the TC-1 cells. In vivo depletion experiments showed that the deactivation of CD80 switched the pro-tumorigenic effect of macrophages observed in TC-1-induced tumors into an anti-tumorigenic effect in TC-1/dCD80-1 tumors and induced the pro-tumorigenic activity of CD4+ cells. Moreover, the frequency of lymphoid and myeloid cells and the CTLA-4 expression by T helper (Th)17 cells were increased in TC-1/dCD80-1- compared with that in the TC-1-induced tumors. CTLA-4 blockade downregulated the frequencies of most immune cell types and upregulated the frequency of M2 macrophages in the TC-1 tumors, while it increased the frequency of lymphoid cells in TC-1/dCD80-1-induced tumors. Furthermore, the anti-CTLA-4 therapy enhanced the frequency of CD8+ T cells as well as CD4+ T cells, especially for a Th1 subset. Regulatory T cells (Treg) formed the most abundant CD4+ T cell subset in untreated tumors. The anti-CTLA-4 treatment downregulated the frequency of Treg cells with limited immunosuppressive potential in the TC-1 tumors, whereas it enriched this type of Treg cells and decreased the Treg cells with high immunosuppressive potential in TC-1/dCD80-1-induced tumors. The immunosuppressive role of tumor-cell-expressed CD80 should be considered in research into biomarkers for the prediction of cancer patients' sensitivity to immune checkpoint inhibitors and for the development of a tumor-cell-specific CD80 blockade.

• Publikační typ
• časopisecké články MeSH

BACKGROUND AND PURPOSE: Src homology 3-domain growth factor receptor-bound 2-like endophilin interacting protein 1 (SGIP1) interacts with cannabinoid CB1 receptors. SGIP1 is abundantly and principally expressed within the nervous system. SGIP1 and CB1 receptors co-localize in axons and presynaptic boutons. SGIP1 interferes with the internalization of activated CB1 receptors in transfected heterologous cells. Consequently, the transient association of CB1 receptors with β-arrestin2 is enhanced and prolonged, and CB1 receptor-mediated ERK1/2 signalling is decreased. Because of these actions, SGIP1 may modulate affect, anxiety, pain processing, and other physiological processes controlled by the endocannabinoid system (ECS). EXPERIMENTAL APPROACH: Using a battery of behavioural tests, we investigated the consequences of SGIP1 deletion in tasks regulated by the ECS in SGIP1 constitutive knockout (SGIP1-/- ) mice. KEY RESULTS: In SGIP1-/- mice, sensorimotor gating, exploratory levels, and working memory are unaltered. SGIP1-/- mice have decreased anxiety-like behaviours. Fear extinction to tone is facilitated in SGIP1-/- females. Several cannabinoid tetrad behaviours are altered in the absence of SGIP1. SGIP1-/- males exhibit abnormal behaviours on Δ9 -tetrahydrocannabinol withdrawal. SGIP1 deletion also reduces acute nociception, and SGIP1-/- mice are more sensitive to analgesics. CONCLUSION AND IMPLICATIONS: SGIP1 was detected as a novel protein associated with CB1 receptors, and profoundly modified CB1 receptor signalling. Genetic deletion of SGIP1 particularly affected behavioural tests of mood-related assessment and the cannabinoid tetrad. SGIP1-/- mice exhibit decreased nociception and augmented responses to CB1 receptor agonists and morphine. These in vivo findings suggest that SGIP1 is a novel modulator of CB1 receptor-mediated behaviour.

• MeSH
• adaptorové proteiny signální transdukční fyziologie   MeSH
• afekt MeSH
• emoce MeSH
• extinkce (psychologie) MeSH
• kanabinoidy MeSH
• myši knockoutované MeSH
• myši MeSH
• nocicepce * MeSH
• receptor kanabinoidní CB1 * genetika   MeSH
• receptory kanabinoidní MeSH
• strach MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH

Programmed cell death protein 1 (PD-1)/PD-1 ligand 1 (PD-L1) blockade is a promising therapy for various cancer types, but most patients are still resistant. Therefore, a larger number of predictive biomarkers is necessary. In this study, we assessed whether a loss-of-function mutation of the interferon (IFN)-γ receptor 1 (IFNGR1) in tumor cells can interfere with anti-PD-L1 therapy. For this purpose, we used the mouse oncogenic TC-1 cell line expressing PD-L1 and major histocompatibility complex class I (MHC-I) molecules and its TC-1/A9 clone with reversibly downregulated PD-L1 and MHC-I expression. Using the CRISPR/Cas9 system, we generated cells with deactivated IFNGR1 (TC-1/dIfngr1 and TC-1/A9/dIfngr1). In tumors, IFNGR1 deactivation did not lead to PD-L1 or MHC-I reduction on tumor cells. From potential inducers, mainly IFN-α and IFN-β enhanced PD-L1 and MHC-I expression on TC-1/dIfngr1 and TC-1/A9/dIfngr1 cells in vitro. Neutralization of the IFN-α/IFN-β receptor confirmed the effect of these cytokines in vivo. Combined immunotherapy with PD-L1 blockade and DNA vaccination showed that IFNGR1 deactivation did not reduce tumor sensitivity to anti-PD-L1. Thus, the impairment of IFN-γ signaling may not be sufficient for PD-L1 and MHC-I reduction on tumor cells and resistance to PD-L1 blockade, and thus should not be used as a single predictive marker for anti-PD-1/PD-L1 cancer therapy.

• MeSH
• antigeny CD274 antagonisté a inhibitory   MeSH
• antigeny CD279 antagonisté a inhibitory   MeSH
• experimentální nádory farmakoterapie   imunologie   metabolismus   patologie   MeSH
• imunoterapie MeSH
• interferon gama antagonisté a inhibitory   MeSH
• myši inbrední C57BL MeSH
• myši MeSH
• nádorové buňky kultivované MeSH
• protinádorové látky imunologicky aktivní farmakologie   MeSH
• transformované buněčné linie účinky léků   imunologie   metabolismus   patologie   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

Regulation of axon guidance and pruning of inappropriate synapses by class 3 semaphorins are key to the development of neural circuits. Collapsin response mediator protein 2 (CRMP2) has been shown to regulate axon guidance by mediating semaphorin 3A (Sema3A) signaling; however, nothing is known about its role in synapse pruning. Here, using newly generated crmp2-/- mice we demonstrate that CRMP2 has a moderate effect on Sema3A-dependent axon guidance in vivo, and its deficiency leads to a mild defect in axon guidance in peripheral nerves and the corpus callosum. Surprisingly, crmp2-/- mice display prominent defects in stereotyped axon pruning in hippocampus and visual cortex and altered dendritic spine remodeling, which is consistent with impaired Sema3F signaling and with models of autism spectrum disorder (ASD). We demonstrate that CRMP2 mediates Sema3F signaling in primary neurons and that crmp2-/- mice display ASD-related social behavior changes in the early postnatal period as well as in adults. Together, we demonstrate that CRMP2 mediates Sema3F-dependent synapse pruning and its dysfunction shares histological and behavioral features of ASD.

• MeSH
• dendritické trny MeSH
• membránové proteiny fyziologie   MeSH
• mezibuněčné signální peptidy a proteiny genetika   MeSH
• myši knockoutované MeSH
• myši MeSH
• neurony MeSH
• neuroplasticita MeSH
• poruchy autistického spektra * MeSH
• proteiny nervové tkáně genetika   fyziologie   MeSH
• semaforiny * MeSH
• signální transdukce MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH