Molecular mechanisms and genetic factors contributing to the developmental dysplasia of the hip.

The most prevalent hip disease in neonates is developmental dysplasia of the hip (DDH). A timely and accurate diagnosis is required to provide the most effective treatment for pediatric patients with DDH. Heredity and gene variation have been the subject of increased attention and research worldwide as one of the factors contributing to the pathogenesis of DDH. Genome-wide association studies (GWAS), genome-wide linkage analyses (GWLA), and exome sequencing (ES) have identified variants in numerous genes and single-nucleotide polymorphisms (SNPs) as being associated with susceptibility to DDH in sporadic and DDH family patients. Furthermore, the DDH phenotype can be observed in animal models that exhibit susceptibility genes or loci, including variants in CX3CR1, KANSL1, and GDF5. The dentification of noncoding RNAs and de novo gene variants in patients with DDH-related syndrome has enhanced our understanding of the genes implicated in DDH. This article reviews the most recent molecular mechanisms and genetic factors that contribute to DDH.

CX3C chemokine: Hallmarks of fibrosis and ageing.

Fibrosis refers to the progressive tissue lesion process characterized by excessive secretion and deposition of extracellular matrix (ECM). Abnormal fibrous tissue deposition distorts tissue architecture and leads to the progressive loss of organ function. Notably, fibrosis is one of the primary pathological appearances of many end stage illnesses, and is considered as a lethal threat to human health, especially in the elderly with ageing-related diseases. CX3C ligand 1 (CX3CL1) is the only member of chemokine CX3C and binds specifically to CX3C receptor 1 (CX3CR1). Different from other chemokines, CX3CL1 possesses both chemotactic and adhesive activity. CX3CL1/CX3CR1 axis involves in various physiological and pathological processes, and exerts a critical role in cells from the immune system, vascular system, and nervous system etc. Notably, increasing evidence has demonstrated that CX3CL1/CX3CR1 signaling pathway is closely related to the pathological process of fibrosis in multiple tissue and organs. We reviewed the crucial role of CX3CL1/CX3CR1 axis in fibrosis and ageing and systematically summarized the underlying mechanism, which offers prospective strategies of targeting CX3C for the therapy of fibrosis and ageing-related diseases.

Sugar and arginine facilitate oral tolerance by ensuring the functionality of tolerogenic immune cell subsets in the intestine.

Although oral tolerance is a critical system in regulating allergic disorders, the mechanisms by which dietary factors regulate the induction and maintenance of oral tolerance remain unclear. To address this, we explored the differentiation and function of various immune cells in the intestinal immune system under fasting and ad libitum-fed conditions before oral ovalbumin (OVA) administration. Fasting mitigated OVA-specific Treg expansion, which is essential for oral tolerance induction. This abnormality mainly resulted from functional defects in the CX3CR1+ cells responsible for the uptake of luminal OVA and reduction of tolerogenic CD103+ dendritic cells. Eventually, fasting impaired the preventive effect of oral OVA administration on asthma and allergic rhinitis development. Specific food ingredients, namely carbohydrates and arginine, were indispensable for oral tolerance induction by activating glycolysis and mTOR signaling. Overall, prior food intake and nutritional signals are critical for maintaining immune homeostasis by inducing tolerance to ingested food antigens.

Redefining the ontogeny of hyalocytes as yolk sac-derived tissue-resident macrophages of the vitreous body.

BACKGROUND: The eye is a highly specialized sensory organ which encompasses the retina as a part of the central nervous system, but also non-neural compartments such as the transparent vitreous body ensuring stability of the eye globe and a clear optical axis. Hyalocytes are the tissue-resident macrophages of the vitreous body and are considered to play pivotal roles in health and diseases of the vitreoretinal interface, such as proliferative vitreoretinopathy or diabetic retinopathy. However, in contrast to other ocular macrophages, their embryonic origin as well as the extent to which these myeloid cells might be replenished by circulating monocytes remains elusive. RESULTS: In this study, we combine transgenic reporter mice, embryonic and adult fate mapping approaches as well as parabiosis experiments with multicolor immunofluorescence labeling and confocal laser-scanning microscopy to comprehensively characterize the murine hyalocyte population throughout development and in adulthood. We found that murine hyalocytes express numerous well-known myeloid cell markers, but concomitantly display a distinct immunophenotype that sets them apart from retinal microglia. Embryonic pulse labeling revealed a yolk sac-derived origin of murine hyalocytes, whose precursors seed the developing eye prenatally. Finally, postnatal labeling and parabiosis established the longevity of hyalocytes which rely on Colony Stimulating Factor 1 Receptor (CSF1R) signaling for their maintenance, independent of blood-derived monocytes. CONCLUSION: Our study identifies hyalocytes as long-living progeny of the yolk sac hematopoiesis and highlights their role as integral members of the innate immune system of the eye. As a consequence of their longevity, immunosenescence processes may culminate in hyalocyte dysfunction, thereby contributing to the development of vitreoretinal diseases. Therefore, myeloid cell-targeted therapies that convey their effects through the modification of hyalocyte properties may represent an interesting approach to alleviate the burden imposed by diseases of the vitreoretinal interface.

Chemokine CX3CL1 (Fractalkine) Signaling and Diabetic Encephalopathy.

Diabetes mellitus (DM) is the most common metabolic disease in humans, and its prevalence is increasing worldwide in parallel with the obesity pandemic. A lack of insulin or insulin resistance, and consequently hyperglycemia, leads to many systemic disorders, among which diabetic encephalopathy (DE) is a long-term complication of the central nervous system (CNS), characterized by cognitive impairment and motor dysfunctions. The role of oxidative stress and neuroinflammation in the pathomechanism of DE has been proven. Fractalkine (CX3CL1) has unique properties as an adhesion molecule and chemoattractant, and by acting on its only receptor, CX3CR1, it regulates the activity of microglia in physiological states and neuroinflammation. Depending on the clinical context, CX3CL1-CX3CR1 signaling may have neuroprotective effects by inhibiting the inflammatory process in microglia or, conversely, maintaining/intensifying inflammation and neurotoxicity. This review discusses the evidence supporting that the CX3CL1-CX3CR1 pair is neuroprotective and other evidence that it is neurotoxic. Therefore, interrupting the vicious cycle within neuron-microglia interactions by promoting neuroprotective effects or inhibiting the neurotoxic effects of the CX3CL1-CX3CR1 signaling axis may be a therapeutic goal in DE by limiting the inflammatory response. However, the optimal approach to prevent DE is simply tight glycemic control, because the elimination of dysglycemic states in the CNS abolishes the fundamental mechanisms that induce this vicious cycle.

Differential alterations of CXCR3, CXCR5 and CX3CR1 in patients with immune thrombocytopenia.

As a versatile element for maintaining homeostasis, the chemokine system has been reported to be implicated in the pathogenesis of immune thrombocytopenia (ITP). However, research pertaining to chemokine receptors and related ligands in adult ITP is still limited. The states of several typical chemokine receptors and cognate ligands in the circulation were comparatively assessed through various methodologies. Multiple variable analyses of correlation matrixes were conducted to characterize the correlation signatures of various chemokine receptors or candidate ligands with platelet counts. Our data illustrated a significant decrease in relative CXCR3 expression and elevated plasma levels of CXCL4, 9-11, 13, and CCL3 chemokines in ITP patients with varied platelet counts. Flow cytometry assays revealed eminently diminished CXCR3 levels on T and B lymphocytes and increased CXCR5 on cytotoxic T cell (Tc) subsets in ITP patients with certain platelet counts. Meanwhile, circulating CX3CR1 levels were markedly higher on T cells with a concomitant increase in plasma CX3CL1 level in ITP patients, highlighting the importance of aberrant alterations of the CX3CR1-CX3CL1 axis in ITP pathogenesis. Spearman's correlation analyses revealed a strong positive association of peripheral CXCL4 mRNA level, and negative correlations of plasma CXCL4 concentration and certain chemokine receptors with platelet counts, which might serve as a potential biomarker of platelet destruction in ITP development. Overall, these results indicate that the differential expression patterns and distinct activation states of peripheral chemokine network, and the subsequent expansion of circulating CXCR5+ Tc cells and CX3CR1+ T cells, may be a hallmark during ITP progression, which ultimately contributes to thrombocytopenia in ITP patients.

Microglial phagocytosis of single dying oligodendrocytes is mediated by CX3CR1 but not MERTK.

Oligodendrocyte death is common in aging and neurodegenerative disease. In these conditions, dying oligodendrocytes must be efficiently removed to allow remyelination and to prevent a feedforward degenerative cascade. Removal of this cellular debris is thought to primarily be carried out by resident microglia. To investigate the cellular dynamics underlying how microglia do this, we use a single-cell cortical demyelination model combined with longitudinal intravital imaging of dual-labeled transgenic mice. Following phagocytosis, single microglia clear the targeted oligodendrocyte and its myelin sheaths in one day via a precise, rapid, and stereotyped sequence. Deletion of the fractalkine receptor, CX3CR1, delays the microglial phagocytosis of the cell soma but has no effect on clearance of myelin sheaths. Unexpectedly, deletion of the phosphatidylserine receptor, MERTK, has no effect on oligodendrocyte or myelin sheath clearance. Thus, separate molecular signals are used to detect, engage, and clear distinct sub-compartments of dying oligodendrocytes to maintain tissue homeostasis.

Treadmill Exercise Enhances Post-Stroke Functional Recovery in Mice via the CX3CL1/CX3CR1 Signaling Pathway.

To validate that treadmill exercise promotes neurofunctional recovery post ischemic stroke and to specifically explore the role of the CX3CL1/CX3CR1 signaling pathway in this treadmill-mediated recovery process. C57BL/6 J mice were used to establish a middle cerebral artery occlusion (MCAO) model. From days 5 to 28 post-stroke, the experimental group did 10-min treadmill sessions twice daily at 12 r/min; the control group remained inactive. On day 6 post-stroke, mice received three intraperitoneal injections of Bromodeoxyuridine (BrdU) or PBS. On days 1, 3, and 5 post-stroke, mice received intracerebroventricular injections of exogenous recombinant CX3CL1, CX3CL1 antagonist, or PBS. The modified neurological severity score (mNSS) and the corner test were used to assess sensorimotor function, and the morris water maze (MWM) test was employed to evaluate cognitive function. Western blot detected CX3CL1 and CX3CR1 protein expression, while immunofluorescence observed these proteins, neurogenesis in the subventricular zone (SVZ), rostral migratory stream (RMS), and dentate gyrus (DG), along with Iba1 and CD68 co-expression. ELISA quantified IL-1ß, IL-4, and IL-10 levels. Treadmill exercise significantly improved neurofunctional recovery in MCAO mice, enhanced neurogenesis in the RMS and SVZ, and increased the expression of CX3CL1 and CX3CR1. The CX3CL1/CX3CR1 axis enhanced the impact of treadmill exercise on neurofunctional recovery, promoting neurogenesis in the RMS and SVZ, and reducing inflammation. Additionally, this axis also enhanced neurogenesis and suppressed microglial activation in the DG induced by treadmill exercise. This study demonstrates the CX3CL1/CX3CR1 pathway as critical for treadmill-induced post-stroke recovery, indicating its potential target for exercise mimetics in rehabilitation.

Induction of CX3CL1 expression by LPS and its impact on invasion and migration in oral squamous cell carcinoma.

Purpose: This study aimed to explore the expression of CX3CL1 induced by lipopolysaccharide (LPS) in oral squamous cell carcinoma (OSCC) and its impact on biological characteristics such as invasion and migration, taking the foundation for new targets for the treatment and prognosis of OSCC. Methods: This study utilized a variety of techniques, including bioinformatics, molecular biology, and cell experiments, to investigate the expression of CX3CL1 and its receptor CX3CR1 in OSCC patients' cancer tissues or OSCC cell lines. Extracting, organizing, and analyzing the TCGA database on the expression of CX3CL1 and its receptor CX3CR1 in cancer tissues and corresponding paracancerous normal tissues of OSCC patients by bioinformatics methods. The expression of CX3CL1 in cancerous and normal tissues of OSCC patients was verified by IHC, and the changes in mRNA and protein expression of CX3CL1 and its receptor CX3CR1 in OSCC cell lines were detected before and after lipopolysaccharide LPS stimulation by RT-PCR, ELISA, and WB. Changes in cell biological behavior by overexpression of CX3CL1 in OSCC cell lines were detected by CCK-8, Transwell, scratch healing assay, and cloning assay. The effects of overexpressing cell lines on the AKT pathway and Epithelial-mesenchymal Transition (EMT)-related protein expression before and after LPS stimulation were detected by Western Blot. Results: (1) CX3CL1 and its receptor CX3CR1 were found to be downregulated in OSCC tissues of patients or OSCC cell lines. (2) After LPS stimulation, CX3CL1 gene expression increased in both OSCC cell lines, while CX3CR1 expression remained unchanged. (3) OSCC cell lines overexpressing CX3CL1 showed changes in cell biological characteristics, including decreased proliferation, invasion, migration, and stemness, which were more pronounced after LPS stimulation. (4) Overexpression of CX3CL1 in OSCC cell lines decreased EMT-related protein expression and AKT phosphorylation. On the contrary were promoted by LPS stimulation. Conclusion: CX3CL1 and CX3CR1 are downregulated in OSCC cancer tissues and cell lines compared to adjacent normal tissues and cells. LPS stimulation increases CX3CL1 expression in OSCC cell lines, suggesting that inflammation may induce CX3CL1 expression and that the CX3CL1 gene may play an important role in OSCC progression. Overexpression of CX3CL1 inhibits OSCC cell proliferation, migration, invasion, and stemness, suggesting that CX3CL1 plays a critical role in suppressing OSCC development. CX3CL1 suppresses OSCC invasion and migration by affecting EMT progression and AKT phosphorylation, and partially reverse the process that LPS causes and affects the development of OSCC.

Upregulation of granzyme B and C-X3-C motif receptor 1 in circulating plasmablasts was negatively regulated by Notch signal in patients with systemic lupus erythematosus.

As one molecule related to cytotoxicity, surface expression of C-X3-C motif receptor 1 (CX3CR1) was highly correlated with intracellular granzyme B (GZMB) in NK and cytolytic T cells. However, the expression of CX3CR1 and GZMB in B cells has not been clarified, and their clinical significance in systemic lupus erythematosus (SLE) remains unclear. This study aimed to clarify the changes and clinical significance of peripheral blood B cells expressing GZMB and/or CX3CR1 in SLE. Peripheral blood was collected from 39 SLE patients and 48 healthy controls. We found that GZMB and CX3CR1 expression varied in different B-cell subsets, with plasmablasts possessing the highest positive percentages, consistent with bioinformatics prediction. GZMB+ and CX3CR1+ percentages in circulating B cells and plasmablasts were increased in SLE patients. CX3CR1 was upregulated on B cells after in vitro stimulation. Notch intracellular domain (NICD) expression was significantly decreased in plasmablasts of SLE patients and CX3CR1 in plasmablasts was downregulated with the addition of JAG1. In conclusion, GZMB and CX3CR1 were increased in B cells and in plasmablasts of SLE patients and CX3CR1 was negatively regulated by Notch signal in plasmablasts, which may be involved in SLE pathogenesis.

CX3CL1-CX3CR1 axis protects retinal ganglion cells by inhibiting microglia activation in a distal optic nerve trauma model.

BACKGROUND: The chemokine CX3CL1 has been reported to play an important role in optic nerve protection, but the underlying mechanism is still unclear. CX3CR1, the only receptor of CX3CL1, is specifically expressed on retinal microglia, whose activation plays a role in the pathological process of optic nerve injury. This study aimed to evaluate whether CX3CL1 exerts optic neuroprotection by affecting the activation of microglia by combining with CX3CR1. METHODS: A mouse model of distal optic nerve trauma (ONT) was used to evaluate the effects of the CX3CL1-CX3CR1 axis on the activation of microglia and survival or axonal regeneration of retinal ganglion cells (RGCs). The activation of microglia, loss of RGCs, and damage to visual function were detected weekly till 4 weeks after modeling. CX3CL1 was injected intravitreally immediately or delayed after injury and the status of microglia and RGCs were examined. RESULTS: Increases in microglia activation and optic nerve damage were accompanied by a reduced production of the CX3CL1-CX3CR1 axis after the distal ONT modeling. Both immediate and delayed intravitreal injection of CX3CL1 inhibited microglia activation, promoted survival of RGCs, and improved axonal regenerative capacity. Injection with CX3CL1 was no longer effective after 48 h post ONT. The CX3CL1-CX3CR1 axis promotes survival and axonal regeneration, as indicated by GAP43 protein and gene expression, of RGCs by inhibiting the microglial activation after ONT. CONCLUSIONS: The CX3CL1-CX3CR1 axis could promote survival and axonal regeneration of RGCs by inhibiting the microglial activation after optic nerve injury. The CX3CL1-CX3CR1 axis may become a potential target for the treatment of optic nerve injury. Forty-eight hours is the longest time window for effective treatment after injury. The study is expected to provide new ideas for the development of targeted drugs for the repair of optic nerve.

m6A Reader YTHDC1 Impairs Respiratory Syncytial Virus Infection by Downregulating Membrane CX3CR1 Expression.

Respiratory syncytial virus (RSV) is the most prevalent cause of acute lower respiratory infection in young children. Currently, the first RSV vaccines are approved by the FDA. Recently, N6-methyladenosine (m6A) RNA methylation has been implicated in the regulation of the viral life cycle and replication of many viruses, including RSV. m6A methylation of RSV RNA has been demonstrated to promote replication and prevent anti-viral immune responses by the host. Whether m6A is also involved in viral entry and whether m6A can also affect RSV infection via different mechanisms than methylation of viral RNA is poorly understood. Here, we identify m6A reader YTH domain-containing protein 1 (YTHDC1) as a novel negative regulator of RSV infection. We demonstrate that YTHDC1 abrogates RSV infection by reducing the expression of RSV entry receptor CX3C motif chemokine receptor 1 (CX3CR1) on the cell surface of lung epithelial cells. Altogether, these data reveal a novel role for m6A methylation and YTHDC1 in the viral entry of RSV. These findings may contribute to the development of novel treatment options to control RSV infection.

Peripheral CX3CR1+ T cells combined with PD-1 blockade therapy potentiates the anti-tumor efficacy for lung cancer.

Identifying tumor-relevant T cell subsets in the peripheral blood (PB) has become a potential strategy for cancer treatment. However, the subset of PB that could be used to treat cancer remains poorly defined. Here, we found that the CX3CR1+ T cell subset in the blood of patients with lung cancer exhibited effector properties and had a higher TCR matching ratio with tumor-infiltrating lymphocytes (TILs) compared to CX3CR1- T cells, as determined by paired single-cell RNA and TCR sequencing. Meanwhile, the anti-tumor activities, effector cytokine production, and mitochondrial function were enhanced in CX3CR1+ T cells both in vitro and in vivo. However, in the co-culture system of H322 cells with T cells, the percentages of apoptotic cells and Fas were substantially higher in CX3CR1+ T cells than those in CX3CR1- T cells. Fas-mediated apoptosis was rescued by treatment with an anti-PD-1 antibody. Accordingly, the combination of adoptive transfer of CX3CR1+ T cells and anti-PD-1 treatment considerably decreased Fas expression and improved the survival of lung xenograft mice. Moreover, an increased frequency of CX3CR1+ T cells in the PB correlated with a better response and prolonged survival of patients with lung cancer who received anti-PD-1 therapy. These findings indicate the promising potential of adoptive transfer of peripheral CX3CR1+ T cells as an individual cancer immunotherapy.

CX3CL1 (Fractalkine)-CX3CR1 Axis in Inflammation-Induced Angiogenesis and Tumorigenesis.

The chemotactic cytokine fractalkine (FKN, chemokine CX3CL1) has unique properties resulting from the combination of chemoattractants and adhesion molecules. The soluble form (sFKN) has chemotactic properties and strongly attracts T cells and monocytes. The membrane-bound form (mFKN) facilitates diapedesis and is responsible for cell-to-cell adhesion, especially by promoting the strong adhesion of leukocytes (monocytes) to activated endothelial cells with the subsequent formation of an extracellular matrix and angiogenesis. FKN signaling occurs via CX3CR1, which is the only known member of the CX3C chemokine receptor subfamily. Signaling within the FKN-CX3CR1 axis plays an important role in many processes related to inflammation and the immune response, which often occur simultaneously and overlap. FKN is strongly upregulated by hypoxia and/or inflammation-induced inflammatory cytokine release, and it may act locally as a key angiogenic factor in the highly hypoxic tumor microenvironment. The importance of the FKN/CX3CR1 signaling pathway in tumorigenesis and cancer metastasis results from its influence on cell adhesion, apoptosis, and cell migration. This review presents the role of the FKN signaling pathway in the context of angiogenesis in inflammation and cancer. The mechanisms determining the pro- or anti-tumor effects are presented, which are the cause of the seemingly contradictory results that create confusion regarding the therapeutic goals.

Discrepant Phenotyping of Monocytes Based on CX3CR1 and CCR2 Using Fluorescent Reporters and Antibodies.

Monocytes, as well as downstream macrophages and dendritic cells, are essential players in the immune system, fulfilling key roles in homeostasis as well as in inflammatory conditions. Conventionally, driven by studies on reporter models, mouse monocytes are categorized into a classical and a non-classical subset based on their inversely correlated surface expression of Ly6C/CCR2 and CX3CR1. Here, we aimed to challenge this concept by antibody staining and reporter mouse models. Therefore, we took advantage of Cx3cr1GFP and Ccr2RFP reporter mice, in which the respective gene was replaced by a fluorescent reporter protein gene. We analyzed the expression of CX3CR1 and CCR2 by flow cytometry using several validated fluorochrome-coupled antibodies and compared them with the reporter gene signal in these reporter mouse strains. Although we were able to validate the specificity of the fluorochrome-coupled flow cytometry antibodies, mouse Ly6Chigh classical and Ly6Clow non-classical monocytes showed no differences in CX3CR1 expression levels in the peripheral blood and spleen when stained with these antibodies. On the contrary, in Cx3cr1GFP reporter mice, we were able to reproduce the inverse correlation of the CX3CR1 reporter gene signal and Ly6C surface expression. Furthermore, differential CCR2 surface expression correlating with the expression of Ly6C was observed by antibody staining, but not in Ccr2RFP reporter mice. In conclusion, our data suggest that phenotyping strategies for mouse monocyte subsets should be carefully selected. In accordance with the literature, the suitability of CX3CR1 antibody staining is limited, whereas for CCR2, caution should be applied when using reporter mice.

Corrigendum: Case Series: Maraviroc and pravastatin as a therapeutic option to treat long COVID/Post-acute sequalae of COVID (PASC).

[This corrects the article DOI: 10.3389/fmed.2023.1122529.].

Decreased exercise-induced natural killer cell redistribution in multiple sclerosis.

BACKGROUND: Exercise may have beneficial effects in MS, remaining controversial its possible disease-modifying effects and which mechanisms might be involved. We evaluated whether exercise-induced lymphocyte redistribution differ in MS patients as compared to controls. METHODS: Exercise was assessed in 12 relapsing-remitting MS patients and 11 controls in a cycle ergometer, obtaining blood samples before exercise, at maximal exercise capacity (T1), and after resting (T2). Peripheral lymphocytes were evaluated by flow cytometry, assessing chemokine receptor expression to study cell trafficking properties. RESULTS: Lymphocyte subsets in all cases increased after exercise and decreased at resting. However, total natural killer (NK) cells in patients as compared to controls had a lower exercise-induced redeployment at T1 (696 ± 581 cells/µL vs.1502 ± 641 cells/µL, p < 0.01). Evaluating NK cell subsets, CD56bright NK cells numbers decreased in peripheral blood in MS patients after resting (T2), contrasting with values remaining above baseline in healthy controls. NK cells mobilized after exercise at T1 in controls, as compared to patients, had a higher CX3CR1 expression (1402 ± 564/µL vs. 615 ± 548 cell//µL, p < 0.01). CONCLUSION: Exercise-induced redeployment of NK cells may be reduced in MS patients, as well as their migration capabilities, pointing to potential immunological mechanisms to be enhanced by exercise training programs.

Single-cell transcriptomics reveal distinct immune-infiltrating phenotypes and macrophage-tumor interaction axes among different lineages of pituitary neuroendocrine tumors.

BACKGROUND: Pituitary neuroendocrine tumors (PitNETs) are common gland neoplasms demonstrating distinctive transcription factors. Although the role of immune cells in PitNETs has been widely recognized, the precise immunological environment and its control over tumor cells are poorly understood. METHODS: The heterogeneity, spatial distribution, and clinical significance of macrophages in PitNETs were analyzed using single-cell RNA sequencing (scRNA-seq), bulk RNA-seq, spatial transcriptomics, immunohistochemistry, and multiplexed quantitative immunofluorescence (QIF). Cell viability, cell apoptosis assays, and in vivo subcutaneous xenograft experiments have confirmed that INHBA-ACVR1B influences the process of tumor cell apoptosis. RESULTS: The present study evaluated scRNA-seq data from 23 PitNET samples categorized into 3 primary lineages. The objective was to explore the diversity of tumors and the composition of immune cells across these lineages. Analyzed data from scRNA-seq and 365 bulk RNA sequencing samples conducted in-house revealed the presence of three unique subtypes of tumor immune microenvironment (TIME) in PitNETs. These subtypes were characterized by varying levels of immune infiltration, ranging from low to intermediate to high. In addition, the NR5A1 lineage is primarily associated with the subtype characterized by limited infiltration of immune cells. Tumor-associated macrophages (TAMs) expressing CX3CR1+, C1Q+, and GPNMB+ showed enhanced contact with tumor cells expressing NR5A1 + , TBX19+, and POU1F1+, respectively. This emphasizes the distinct interaction axes between TAMs and tumor cells based on their lineage. Moreover, the connection between CX3CR1+ macrophages and tumor cells via INHBA-ACVR1B regulates tumor cell apoptosis. CONCLUSIONS: In summary, the different subtypes of TIME and the interaction between TAM and tumor cells offer valuable insights into the control of TIME that affects the development of PitNET. These findings can be utilized as prospective targets for therapeutic interventions.

Triptolide alleviates cerebral ischemia/reperfusion injury via regulating the Fractalkine/CX3CR1 signaling pathway.

PURPOSE: To evaluate the potential efficacy of Triptolide (TP) on cerebral ischemia/reperfusion injury (CIRI) and to uncover the underlying mechanism through which TP regulates CIRI. METHODS: We constructed a middle cerebral artery occlusion/reperfusion (MCAO/R) mouse model to simulate CIRI, and established a lipopolysaccharide (LPS)-stimulated BV-2 cell model to mimic the inflammatory state during CIRI. The neurological deficits score (NS) of mice were measured for assessment of neurologic functions. Both the severity of cerebral infarction and the apoptosis level in mouse brain tissues or cells were respectively evaluated using corresponding techniques. The expression levels of Ionized calcium binding adapter molecule 1 (IBA-1), Inductible Nitric Oxide Synthase (iNOS), Arginase 1 (Arg-1), Tumor necrosis factor-α (TNF-α), Interleukin 1ß (IL-1ß), Cysteine histoproteinase S (CTSS), Fractalkine, chemokine C-X3-C motif receptor 1 (CX3CR1), BCL-2-associated X protein (BAX), and antiapoptotic proteins (Bcl-2) were detected using immunofluorescence, qRT-PCR as well as Western blot, respectively. RESULTS: Relative to the Sham group, treatment with TP attenuated the increased NS, infarct area and apoptosis levels observed in MCAO/R mice. Upregulated expression levels of IBA-1, iNOS, Arg-1, TNF-α and IL-1ß were found in MCAO/R mice, while TP suppressed iNOS, TNF-α and IL-1ß expression, and enhanced Arg-1 expression in both MCAO/R mice and LPS-stimulated BV-2 cells. Besides, TP inhibited the CTSS/Fractalkine/CX3CR1 pathway activation in both MCAO/R mice and LPS-induced BV-2 cells, while overexpression of CTSS reversed such effect. Co-culturing HT-22 cells with TP+LPS-treated BV-2 cells led to enhanced cell viability and decreased apoptosis levels. However, overexpression of CTSS further aggravated HT-22 cell injury. CONCLUSION: TP inhibits not only microglia polarization towards the M1 phenotype by suppressing the CTSS/Fractalkine/CX3CR1 pathway activation, but also HT-22 apoptosis by crosstalk with BV-2 cells, thereby ameliorating CIRI. These findings reveal a novel mechanism of TP in improving CIRI, and offer potential implications for addressing the preventive and therapeutic strategies of CIRI.