BACKGROUND: Early-onset schizophrenia (EOS) is a type of schizophrenia (SCZ) with an age of onset of < 18 years. An abnormal inflammatory immune system may be involved in the occurrence and development of SCZ. We aimed to identify the immune characteristic genes and cells involved in EOS and to further explore the pathogenesis of EOS from the perspective of immunology.
METHODS: We obtained microarray data from a whole-genome mRNA expression in peripheral blood mononuclear cells (PBMCs); 19 patients with EOS (age range: 14.79 ± 1.90) and 18 healthy controls (HC) (age range: 15.67 ± 2.40) were involved. We screened for differentially expressed genes (DEGs) using the Limma software package and modular genes using weighted gene co-expression network analysis (WGCNA). In addition, to identify immune characteristic genes and cells, we performed enrichment analysis, immune infiltration analysis, and receiver operating characteristic (ROC) curve analysis; we also used a random forest (RF), a support vector machine (SVM), and the LASSO-Cox algorithm.
RESULTS: We selected the following immune characteristic genes: CCL8, PSMD1, AVPR1B and SEMG1. We employed a RF, a SVM, and the LASSO-Cox algorithm. We identified the following immune characteristic cells: activated mast cells, CD4+ memory resting T cells, resting mast cells, neutrophils and CD4+ memory activated T cells. In addition, the AUC values of the immune characteristic genes and cells were all > 0.7.
CONCLUSIONS: Our results indicate that immune system function is altered in SCZ. In addition, CCL8, PSMD1, AVPR1B and SEMG1 may regulate peripheral immune cells in EOS. Further, immune characteristic genes and cells are expected to be diagnostic markers and therapeutic targets of SCZ.

Tumor-associated macrophages (TAMs) play a pivotal role in reshaping the tumor microenvironment following radiotherapy. The mechanisms underlying this reprogramming process remain to be elucidated.
Subcutaneous Lewis lung carcinoma (LLC) murine model was treated with hypofrationated radiotherapy (8 Gy × 3F). Single-cell RNA sequencing was utilized to identify subclusters and functions of TAMs. Multiplex assay and enzyme-linked immunosorbent assay (ELISA) were employed to measure serum chemokine levels. Bindarit was used to inhibit CCL8, CCL7, and CCL2. The infiltration of TAMs after combination treatment with hypofractionated radiotherapy and Bindarit was quantified with flow cytometry, while the influx of CD206 and CCL8 was assessed by immunostaining.
Transcriptome analysis identified a distinct subset of M2-like macrophages characterized by elevated Ccl8 expression level following hypofractionated radiotherapy in LLC-bearing mice. Remarkbly, hypofractionated radiotherapy not only promoted CCL8high macrophages infiltration but also reprogrammed them by upregulating immunosuppressive genes, thereby fostering an immunosuppressive tumor microenvironment. Additioinally, hypofractionated radiotherapy enhanced the CCL signaling pathway, augmenting the pro-tumorigenic functions of CCL8high macrophages and boosting TAMs recruitment. The adjunctive treatment combining hypofractionated radiotherapy with Bindarit effectively reduced M2 macrophages infiltration and prolonged the duration of local tumor control.
Hypofractionated radiotherapy enhances the infiltration of CCL8high macrophages and amplifies their roles in macrophage recruitment through the CCL signaling pathway, leading to an immunosuppressive tumor microenvironment. These findings highlight the potential of targeting TAMs and introduces a novel combination to improve the efficacy of hypofractionated radiotherapy.

Vitiligo is a T cell-mediated skin depigmentation disease. Though treatments arresting disease progression and inducing repigmentation are available, the efficacy of these options is often limited and poorly sustained. How stromal signals contribute to the interferon-γ-dominant skin niches is unclear. This study aims to determine how fibroblasts participate in the IFN-γ-dominant vitiligo niche.
Mouse vitiligo models were established. Fibroblasts from control and vitiligo mice were extracted for RNA sequencing. In vitro IFN-γ stimulation was performed to verify the JAK-STAT pathway by qPCR and Western blot. T cell polarization with chemokines was measured by flow cytometry. Protein levels in tissues were also examined by IHC.
The vitiligo mouse model recapitulates the human CD8-IFN-γ pathway. RNA sequencing revealed elevated chemokine CCL2 and CCL8 in vitiligo fibroblast, which may be regulated by the JAK-STAT signaling. Such phenomenon is verified by JAK inhibitor peficitinib in vitro. Moreover, CCL2 addition into the naïve T polarization system promoted type 2 cytokines secretion, which represents a hallmark of vitiligo lesions.
Dermal fibroblasts, a principal constituent of skin structure, respond to IFN-γ by skewing T cells towards a type 2 cytokine profile via CCL2 and CCL8, which can be abrogated by JAK inhibitor peficitinib.

Prior investigations of the tumor microenvironment (TME) of diffuse large B-cell lymphoma (DLBCL) have shown that immune and stromal cells are key contributing factors to patients\' outcome. However, challenges remain in finding reliable prognostic biomarkers based on cell infiltration. In this study, we attempted to shed some light on chemokine C-C motif chemokine ligand 8 (CCL8) in DLBCL via interaction with M2 macrophages.
The Estimation of STromal and Immune cells in MAlignant Tumor tissues using Expression data (ESTIMATE) algorithm was applied to evaluate immune and stromal scores from transcriptomic profiles of 443 DLBCL samples from The Cancer Genome Atlas (TCGA) and GSE10846 datasets. Immune cell infiltration (ICI) clusters were obtained based on different immune cell infiltrations of each sample, and gene clusters were derived through differentially expressed genes (DEGs) between the distinct ICI clusters. Five immune-related hub genes related to overall survival (OS) and clinical stages were obtained by COX regression analysis and protein-protein interaction (PPI) network construction then verified by quantitative real-time PCR (qPCR) and immunofluorescence staining in the FFPE tissues. The Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), and TIMER websites were employed to explore the biological functions of CCL8-related DEGs. Uni- and multivariable Cox regression analyses were performed to analyze CCL8 as an independent prognostic risk factor in GSE10846 and were verified in other independent GEO cohorts.
A higher stromal score was associated with favorable prognosis in DLBCL. Patients in the ICI B cluster and gene B clusters had a better follow-up status with a higher programmed death ligand 1 (PD-L1) and cytotoxic T-lymphocyte antigen 4 (CTLA4) expression. Most of ICI-related DEGs were enriched for immune-related signaling pathways. Five hub genes with a distinct prognosis association were identified, including CD163, which is a biomarker of M2 macrophages, and CCL8. Abundant M2 macrophages were discovered in the high-CCL8 expression group. The functional analysis indicated that CCL8 is a key component of immune-related processes and secretory granule groups. Cox regression analysis and data from other GSE datasets yielded additional evidence of the prognostic value of CCL8 in DLBCL.
CCL8 has been implicated in macrophage recruitment in several solid tumors, and only a few reports have been published on the role of CCL8 in the pathogenesis of hematological malignancies. This article attempted to find out TME-related genes that associated with the survival in DLBCL patients. CCL8 was identified to be involved in immune activities. Importantly, a series of bioinformatics analysis indicated that CCL8 might become an effective target for DLBCL, which interacts with M2 macrophage and immune checkpoint. The potential related mechanisms need to be further elucidated.

What is the central question of this study? What is the relationship of chemokine (C-C motif) ligand 8 (CCL8) to thoracic aortic aneurysm and dissection (TAAD) formation in postnatal mice with vascular smooth muscle cell (VSMC) Tgfbr2 disruption, and is dexamethasone a potential treatment? What is the main finding and its importance? CCL8 was associated with the formation of TAAD in VSMC Tgfbr2-disrupted mice. Dexamethasone reduced TAAD formation and inhibited mitogen-activated protein kinase (p-p38) and nuclear factor-κB (p-p65) signalling pathways. CCL8 might be an important promoter of aortic inflammation. Dexamethasone provided potential therapeutic effects in TAAD treatment.
Aortic inflammation plays a vital role in initiation and progression of thoracic aortic aneurysm and dissection (TAAD). Disturbance of the transforming growth factor-β (TGF-β) signalling pathway is believed to be one of the pathogenic mechanisms of TAAD. Initially, Myh11-CreERT2 .Tgfbr2f/f male mice were used to build a TAAD mouse model, and bioinformatic analyses revealed enriched inflammatory signal pathways and upregulated chemokine (C-C motif) ligand 8 (CCL8). So we hypothesized that vascular smooth muscle cell (VSMC) Tgfbr2 disruption in postnatal mice results in aortic inflammation associated with CCL8 secretion. Real-time quantitative PCR and serum enzyme-linked immunosorbent assay (ELISA) results confirmed that CCL8 expression began to increase after VSMC Tgfbr2 disruption. Next, we cultured mouse thoracic aortas ex vivo, and observed that the protein expression of CCL8 in culture supernatants was increased by ELISA. Subsequently, the co-localization of CCL8 with α-smooth muscle actin or CD68 was found to be significantly increased by immunofluorescence. Then, dexamethasone (DEX) was used to treat TAAD in VSMC Tgfbr2-disrupted mice; the results of histochemical, immunofluorescence and immunohistochemical staining indicated that DEX therapy reduced CCL8 secretion, inflammatory cell recruitment, aortic medial thickening, elastic fibre fragmentation, extracellular matrix degradation and contractile apparatus impairment, and thereby ameliorated TAAD formation. Western blotting showed that mitogen-activated protein kinase and nuclear factor-κB signalling pathways in aorta were overactivated after VSMC Tgfbr2 disruption, but inhibited by DEX therapy. Altogether, CCL8 might be an important promoter in TAAD formation of VSMC Tgfbr2-disrupted mice, and DEX provided potential therapeutic effects in TAAD treatment.

Chemokines have been reported to play important roles in atherosclerotic development. Recently, we found C-C motif ligand 8 (CCL8), a rarely studied chemokine in atherosclerosis, was highly expressed in the endothelium of advanced human carotid plaques. We hypothesized whether CCL8 promotes atherosclerosis through endothelial dysfunction. Apolipoprotein E-deficient mice under the Western diet were used to construct atherosclerosis models. Adeno-associated viruses (AAV) with CCL8 and the CCL8-antibody were injected into mice respectively to conduct CCL8 overexpression and suppression. The results showed that atherosclerotic lesions were significantly increased in the AAV-CCL8 group, while, lesions in the aortic sinus were reduced in the CCL8-antibody group. With CCL8 treatment (200 ng/ml, 24 h) in vitro, the permeability of human aortic endothelial cells (HAECs) increased and the expression of junctional proteins Zonula occluden-1, and Vascular endothelial cadherin were decreased. This effect was dependent on reactive oxygen species (ROS) generation, which could be blocked by l-Ascorbic acid and Apocynin. Results showed that NADPH oxidase 2 (NOX2) expression also increased with CCL8 stimulation and the ROS, and permeability increase of HAECs could be inhibited when NOX2 interfered with the specific siRNA. Additionally, we further found ERK1/2, PI3K-AKT, and NF-κB pathways were involved in the activation of CCL8. Our results indicated that CCL8 might also play important roles in atherosclerosis and this effect, at least in part, was caused by NOX2/ROS-induced endothelial permeability increase. This study might contribute to a deeper understanding of the connection between chemokines and atherosclerosis.

OBJECTIVE: To investigate the roles of chemokine (C-C motif) ligand 8 (CCL8) in periodontal ligament during orthodontic tooth movement (OTM).
METHODS: Bioinformatics analyzed 100 genes in human periodontal ligament cells that were most upregulated after 48 hours of mechanical stress, and these genes were classified through GO and KEGG databases. Nickel-titanium closed-coil springs were placed between right first molar and incisors to produce 20 cN of orthodontic force in eight-week-old male SD rats for 1 and 2 days, followed by immunohistochemical staining of CCL8. Human periodontal ligament fibroblasts (hPDLFs) were stimulated by 14% cyclic tension force (Flexcell FX-5000 T Tension System) or hypoxia conditions to mimic OTM for 1 and 2 days, then the resulting CCL8 were examined through ELISA. Scratching assay was performed by treating hPDLFs with different concentrations of CCL8 (1 ng/ml, 10 ng/ml, 100 ng/ml). The migration, proliferation, and adhesion abilities of 100 ng/ml CCL8-treated hPDLFs were also examined. qRT-PCR and western blot detected matrix metalloproteinase 3, periostin, and osteoprotegrin expressions of hPDLFs under 100 ng/ml CCL8.
RESULTS: Bioinformatic analysis demonstrated that CCL8 was upregulated after applying mechanical stress for 48 hours. CCL8 secretion showed upregulation after 24 hours of OTM applicationsin vivo and in vitro. CCL8-treated hPDLFs showed significant positive effects on cell proliferation and matrix metalloproteinase 3. It also inhibited periostin and osteoprotegrin expressions.
CONCLUSIONS: CCL8 was upregulated in periodontal ligament during initial stage of OTM. Although CCL8 in human periodontal ligaments showed no significant effects on cell migration ability, it did enhance cell proliferation and osteoclastogenesis.

OBJECTIVE: The theory of inflammation is one of the important theories in the pathogenesis of diabetic nephropathy (DN). We herein aimed to explore whether loganin affected macrophage infiltration and activation upon diabetic nephropathy (DN) by a spontaneous DN mice and a co-culture system of glomerular mesangial cells (GMCs) and macrophage cells (RAW264.7) which was induced by advanced glycation end products (AGEs).
UNASSIGNED: Loganin showed remarkable capacity on protecting renal from damage by mitigating diabetic symptoms, improving the histomorphology of the kidney, decreasing the expression of extracellular matrix such as FN, COL-IV and TGF-β, reversing the production of IL-12 and IL-10 and decreasing the number of infiltrating macrophages in the kidney. Moreover, loganin showed markedly effects by suppressing iNOS and CD16/32 expressions (M1 markers), increasing Arg-1 and CD206 expressions (M2 markers), which were the phenotypic transformation of macrophage. These effects may be attributed to the inhibition of the receptor for AGEs (RAGE) /monocyte chemotactic protein-1 (MCP-1)/CC chemokine receptor 2 (CCR2) signaling pathway, with significantly down-regulated expressions of RAGE, MCP-1 and CCR2 by loganin. Loganin further decreased MCP-1 secretion when RAGE was silenced, which means other target was involved in regulating the MCP-1 expression. While loganin combinated with the inhibitor of CCR2 exerted stronger anti-inhibition effects of iNOS expression, suggesting that CCR2 was the target of loganin in regulating the activation of macrophages.
CONCLUSIONS: Loganin could ameliorate DN kidney damage by inhibiting macrophage infiltration and activation via the MCP-1/CCR2 signaling pathway in DN.

Breast cancer (BC) is the most common malignancy among females worldwide. The tumor microenvironment usually prevents effective lymphocyte activation and infiltration, and suppresses infiltrating effector cells, leading to a failure of the host to reject the tumor. CC chemokines play a significant role in inflammation and infection.
In our study, we analyzed the expression and survival data of CC chemokines in patients with BC using several bioinformatics analyses tools.
The mRNA expression of CCL2/3/4/5/7/8/11/17/19/20/22 was remarkably increased while CCL14/21/23/28 was significantly down-regulated in BC tissues compared with normal tissues. Methylation could down-regulate expression of CCL2/5/15/17/19/20/22/23/24/25/26/27 in BC. Low expression of CCL3/4/23 was found to be associated with drug resistance in BC. Results from Kaplan-Meier plotter and BC Gene-Expression Miner v4.2 (bcGenExMiner) v4.2 demonstrated that BC patients with high CCL8 and low CCL19/21/22 expression were more likely to have a worse prognosis. CCL8 expression was significantly up-regulated in BC tissues compared with normal tissues. High CCL8 expression was significantly correlated with negative PR, negative ER, positive nodal status, triple-negative BC subtype, basal-like BC subtype, triple-negative and basal-like BC subtype and high grades. CCL21 was down-regulated in BC, while high levels of CCL21 was associated with negative PR, triple-negative subtype, basal-like subtype and low tumor grade. Functional analysis demonstrated that CCL8 and CCL21 were involved in carcinogenesis, tumor immune escape and chemoresistance in BC.
Integrative bioinformatics analysis demonstrated CCL8/21 as potential prognostic biomarkers in BC microenvironment.

OBJECTIVE: Angiogenesis is an important progress associated with several pathological situations. Several chemokines have been reported to act as regulators of angiogenesis. The current study aimed to find whether C-C Motif Chemokine 8 is involved in angiogenesis regulation.
METHODS: To verify whether C-C Motif Chemokine 8 is related to angiogenesis in plaques, carotid plaques were collected from patients with severe carotid stenosis and analysed using CD31 immunohistochemistry and real-time PCR. To further clarify the relation between C-C Motif Chemokine 8 and angiogenesis, human umbilical vein endothelium cells and human dermal microvascular endothelial cells were treated with C-C Motif Chemokine 8 in the presence or absence of C-C motif chemokine receptor 2-Ab and extracellular regulated MAP kinase 1/2 inhibition (FR180204). Proliferation and migration of human umbilical vein endothelium cells and human dermal microvascular endothelial cells were examined with Cell Counting Kit-8 and Transwell chamber assay, respectively. In vitro angiogenesis stimulated by C-C Motif Chemokine 8 was examined using tube formation assay. Ex vivo and in vivo angiogenesis were assessed by mice aortic ring assay and Matrigel plug assay, respectively. C-C motif chemokine receptors of human umbilical vein endothelium cells were examined with real-time PCR, and C-C motif chemokine receptor 1, C-C motif chemokine receptor 2, extracellular regulated MAP kinase 1/2 and phosphorylation-extracellular regulated MAP kinase 1/2 were examined with western blotting assay.
RESULTS: C-C Motif Chemokine 8 was increased in carotid plaques with severe angiogenesis in both RNA and protein level. C-C Motif Chemokine 8 (5 ng/ml) weakly increased human umbilical vein endothelium cell proliferation, but not on human dermal microvascular endothelial cells. Migration and tube formation could be induced by C-C Motif Chemokine 8 in both human umbilical vein endothelium cells and human dermal microvascular endothelial cells. In mice aortic ring assay and Matrigel plug assay, C-C Motif Chemokine 8 could promote angiogenesis compared to vehicle groups. Phosphorylation of extracellular regulated MAP kinase 1/2 was increased with C-C Motif Chemokine 8 stimulation. The migration and tube formation promoted by C-C Motif Chemokine 8 could be largely blocked by C-C motif chemokine receptor 2-Ab or extracellular regulated MAP kinase 1/2 inhibition (FR180204).
CONCLUSIONS: C-C Motif Chemokine 8 could promote both in vitro and in vivo angiogenesis. C-C motif chemokine receptor 2 played an important role in the activation of C-C Motif Chemokine 8 and extracellular regulated MAP kinase 1/2 signalling pathway was involved in this mechanism.