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Peritoneal dissemination is a predominant pattern of metastasis in patients with advanced ovarian cancer. Despite recent progress in the management strategy, peritoneal dissemination remains a determinant of poor ovarian cancer prognosis. Using various histological types of patient-derived ovarian cancer organoids, the roles of the apicobasal polarity of ovarian cancer cell clusters in peritoneal dissemination were studied. First, it was found that both ovarian cancer tissues and ovarian organoids showed apicobasal polarity, where zonula occludens-1 (ZO-1) and integrin beta 4 (ITGB4) served as markers for apical and basal sides, respectively. The organoids in suspension culture, as a model of cancer cell cluster floating in ascites, showed apical-out/basal-in polarity status, while once embedded in extracellular matrix (ECM), the organoids switched their polarity to apical-in/basal-out. This polarity switch was accompanied by the SRC kinase family (SFK) phosphorylation and was inhibited by SFK inhibitors. SFK inhibitors abrogated the adherence of the organoids onto the ECM-coated plastic surface. When the organoids were seeded on a mesothelial cell layer, they cleared and invaded mesothelial cells. In vivo, dasatinib, an SFK inhibitor, suppressed peritoneal dissemination of ovarian cancer organoids in immunodeficient mice. These results suggest SFK-mediated polarity switching is involved in peritoneal metastasis. Polarity switching would be a potential therapeutic target for suppressing peritoneal dissemination in ovarian cancer.

Apico-basal polarity is a fundamental property of the epithelium that functions as a barrier, holds cells together, and determines the directions of absorption and secretion. Apico-basal polarity is regulated by extracellular matrix-integrin binding and downstream signaling pathways, including focal adhesion kinase, rouse-sarcoma oncogene (SRC), and RHO/RHO-associated kinase (ROCK). Loss of epithelial cell polarity plays a critical role in the progression of cancer cells. However, in differentiated carcinomas, polarity is not completely lost but dysregulated. Recent progress with a three-dimensional culture of primary cancer cells allowed for studies of the mechanism underlying the abnormality of polarity in differentiated cancers, including flexible switching of polarity status in response to the microenvironment. Invasive micropapillary carcinoma (MPC) is one of the histopathological phenotypes of adenocarcinoma, which is characterized by inverted polarity. Aberrant activation of RHO-ROCK signaling plays a critical role in the MPC phenotype. Establishing in vitro models will contribute to future drug targeting of the abnormal polarity status in cancer.

Bacteria that exhibit polar growth, i.e. build their peptidoglycan cell walls in restricted zones at cell poles, often show large morphological diversity and plasticity. However, their mechanisms for regulation of cell shape and cell wall assembly are poorly understood. The Gram-positive Streptomyces bacteria, like other Actinobacteria, depend on the essential coiled coil protein DivIVA for establishment of cell polarity and direction of polar growth. Streptomycetes grow as filamentous hyphae that exhibit tip extension. New hyphal tips are generated by lateral branching. Cell shape is largely determined by the control of cell wall growth at these hyphal tips. The Ser/Thr protein kinase AfsK is involved in controlling polar growth and directly phosphorylates DivIVA. Here, we identify a protein phosphatase in Streptomyces coelicolor, SppA, that dephosphorylates DivIVA in vivo and in vitro and affects growth and cell shape. An sppA mutant shows reduced rate of hyphal tip extension, altered hyphal branching patterns, and exhibits frequent spontaneous hyphal growth arrests, all contributing to the unusually dense mycelial structure and slow growth rate that characterize sppA mutants. These phenotypes are largely suppressed in an afsK sppA double mutant, showing that AfsK and SppA partially affect the same regulatory pathway and share target proteins that are involved control of polar growth in S. coelicolor. Strains with a non-phosphorylatable mutant DivIVA were constructed and confirm that the effect of afsK on hyphal branching during normal growth is mediated by DivIVA phosphorylation. However, the phenotypic effects of sppA deletion are independent of DivIVA phosphorylation and must be mediated via other substrates. Altogether, this study identifies a PPP-family protein phosphatase directly involved in the control of polar growth and cell shape determination in S. coelicolor and underscore the importance of eukaryotic-type Ser/Thr phosphorylation in regulation of growth and cell envelope biogenesis in Actinobacteria.Author summaryGram-positive bacteria often use eukaryotic-like Ser/Thr protein phosphorylation in the regulation of central processes related to growth and cell cycle progression. Members of the Actinobacteria, including pathogenic mycobacteria and antibiotic-producing streptomycetes, exhibit a distinctive mode of polar growth, with cell wall synthesis being restricted to zones at cell poles and directed by the essential cell polarity determinant DivIVA. Previously, we have shown that Streptomyces coelicolor modulates its machinery for polar growth and cell shape determination via the Ser/Thr kinase AfsK, which phosphorylates DivIVA. Here, we identify a phosphoprotein phosphatase that targets DivIVA and reverses the AfsK-mediated phosphorylation. We show that this phosphatase has strong effects on polar growth and cell shape, including hyphal branching and the rate of tip extension. The effects are to a large extent dependent on the kinase AfsK but do not require DivIVA phosphorylation as such. The results identify a new regulator of polar growth in streptomycetes, and highlight the importance of Ser/Thr phosphorylation in regulation of actinobacterial growth and cell shape determination.

Wnt signaling plays an essential role in the initiation and progression of various types of cancer. Besides, the Wnt pathway components have been established as reliable biomarkers and potential targets for cancer therapy. Wnt signaling is categorized into canonical and noncanonical pathways. The canonical pathway is involved in cell survival, proliferation, differentiation and migration, while the noncanonical pathway regulates cell polarity and migration. Apart from its biological role in development and homeostasis, the Wnt pathway has been implicated in several pathological disorders, including cancer. As a result, inhibiting this pathway has been a focus of cancer research with multiple targetable candidates in development. In this review, our focus will be to summarize information about ongoing and completed clinical trials targeting various Wnt pathway components, along with describing current and emerging Wnt targeted therapies. In addition, we will discuss potential opportunities and associated challenges of inhibiting Wnt signaling for cancer therapy.

External genitalia development in mice involves multiple developmental processes under the regulation of various signaling pathways. Wnt5a, one of the major Wnt ligands, is a crucial developmental regulator of outgrowing organs such as the limb, the mandible, and the external genitalia. Defects in Wnt5a signaling have been linked to Robinow syndrome, a genetic disorder in which male patients manifest a micropenis and defective urethral tube formation. Whereas Wnt5a is required for cell proliferation during embryonic external genitalia outgrowth, its role for urethral tube formation has yet to be understood. Here, we show that Wnt5a contributes to urethral tube formation as well as external genitalia outgrowth. Wnt5a is expressed in the embryonic external genitalia mesenchyme, and mesenchymal-specific conditional Wnt5a knockout mice resulted in hypospadias-like urethral defects. Early deletion of Wnt5a at E10.5 showed severe defects in both external genitalia outgrowth and urethral tube formation, along with reduced cell proliferation. The severe urethral tube defect persisted during later timing deletion of Wnt5a (E13.5). Further analyses revealed that loss of Wnt5a disrupted cell polarity and led to a reduction of the phosphorylated myosin light chain and the focal adhesion protein, vinculin. Altogether, these results suggest that Wnt5a coordinates cell proliferation and directed cell migration in a stage-dependent manner during male external genitalia development. Furthermore, Wnt5a may regulate cell polarity, focal adhesion formation, and cell contractility, leading to directed cell migration during male-type urethral formation in a manner that has not been reported in other organ fusion events.

Macrophage polarization is of great importance in the formation of atherosclerotic plaque. Homeobox A5 (HOXA5), one of the homeobox transcription factors, has been revealed to be closely associated with macrophage phenotype switching. This study aims to investigate the role of HOXA5 in carotid atherosclerosis (CAS). Herein, the role of HOXA5 was explored in polarized RAW264.7 macrophages in vitro and ApoE-/- mice in vivo. Interestingly, compared with that in M0 macrophages, both the mRNA and protein expression levels of HOXA5 were decreased in lipopolysaccharide (LPS)/interferon (IFN)-γ-induced M1 macrophages, while increased in IL-4-induced M2 macrophages. In addition, in the presence of IL-4, HOXA5-overexpressing RAW264.7 cells preferred to polarizing toward M2 phenotypes. Furthermore, we found that HOXA5 bound to the promoter region and activated the expression of mediator subunit 1 (MED1), a gene known to regulate macrophage differentiation. Knocking MED1 down inhibited HOXA5-enhanced M2 macrophage polarization. In vivo, the CAS model was induced in ApoE-/- mouse fed with a Western-type diet and placed a perivascular carotid collar. Decreased mRNA and protein expressions of HOXA5 were observed in carotid arteries of CAS mice. Forced overexpression of HOXA5 reduced intimal hyperplasia and lipid accumulation in carotid vessels, and it also promoted the polarization of macrophages to M2 subtypes. The expression of MED1 was decreased in atherosclerotic carotid vessels, while HOXA5 overexpression restored its change. Collectively, HOXA5 in carotid arteries is involved in the macrophage M1/M2 switching in atherosclerotic plaque, which may be associated with its transcriptional regulation of MED1.

Background Human endogenous retrovirus‐H long terminal repeat‐associating protein 2 (HHLA2) is a member of B7 family, which is upregulated in multiple tumors. However, its exact functions in non‐small cell lung cancer (NSCLC) have not been fully understood. This study aimed to investigate the biological roles of HHLA2 in human NSCLC and the relevant mechanisms. In addition, the effects of tumor cell‐derived HHLA2 on tumor‐associated macrophage (TAM) polarization were explored. Methods NSCLC cell growth, migration, and invasion were assessed by colony formation and modified Boyden chamber assays. Cell cycle and the CD163+ TAMs were examined by flow cytometry. A co‐culture model of THP‐1 macrophages and NSCLC cells was conducted to investigate the impacts of tumor cell‐derived HHLA2 on THP‐1 macrophage polarization. Moreover, a xenograft nude mouse model was established to explore the effects of HHLA2 on tumorigenesis in vivo. Results HHLA2 was upregulated in A549 and H1299 cells compared with the normal lung epithelial BEAS‐2B cells. HHLA2 deficiency inhibited NSCLC cell proliferation, migration, invasion, and induced G0/G1 phase arrest partially via inhibiting EGFR/MAPK/ERK signaling pathway. Furthermore, HHLA2 knockdown inhibited M2 polarization of TAMs via downregulating IL‐10. In addition, knockdown of HHLA2 inhibited tumor growth in vivo. Conclusion HHLA2 downregulation inhibited NSCLC growth and TAM M2 polarization. HHLA2 may serve as a therapeutic target and promising prognostic biomarker in NSCLC., HHLA2 deficiency inhibited the proliferation, migration, invasion, and induced G0/G1 arrest of A549 and H1299 cells by inactivating EGFR/MAPK/ERK signaling pathway. In addition, knockdown of HHLA2 in NSCLC cells inhibited M2 polarization of THP‐1 macrophages via decreasing the secretion of IL‐10.

Dasatinib, a tyrosine kinase inhibitor, has a protective effect on experimental acute respiratory distress syndrome (ARDS). This study investigated the effect and mechanism of dasatinib in ARDS. C57BL/6 mice were administered with dasatinib (1 and 10 mg/kg) after lipopolysaccharide (LPS) treatment to evaluate the effect of dasatinib on white blood cells (WBC), neutrophils, lymphocytes and macrophages in bronchoalveolar lavage fluid (BALF). The levels and mRNA expressions of inflammation-related cytokines in lung tissues and RAW 264.7 cells were detected by enzyme-linked immunosorbent assay and quantitative real-time PCR, respectively. The protein expressions of nuclear factor E2-related factor 2 (Nrf2) and heme oxygenase 1 (HO1) were determined by Western blot. MTT assay was performed to detect the viability of RAW 264.7 cell. Rescue experiments were used to assess the effect of Nrf2 silencing on the LPS- and dasatinib-treated mice. Under LPS treatment, levels of the WBC, neutrophils, lymphocytes and macrophages in BALF and mRNA expressions of IL-6, TNF-α and IL-10 as well as expression of iNOS were increased, but the expression of arginase-1 was inhibited, while no obvious changes of the protein expressions of Nrf2 and HO1 were observed. Dasatinib partially reversed the effects of LPS above, and further promoted the mRNA expression of IL-10 and the protein expressions of Nrf2 and HO1, while Nrf2 silencing counteracted the effect of dasatinib. Dasatinib induced the polarization of M2 subtype of macrophages and alleviated LPS-induced ARDS through activating Nrf2 signaling pathway, which may provide a new strategy for the treatment of ARDS.

Alternative polarization of macrophages regulates multiple biological processes. While M1‐polarized macrophages generally mediate rapid immune responses, M2‐polarized macrophages induce chronic and mild immune responses. In either case, polyunsaturated fatty acid (PUFA)‐derived lipid mediators act as both products and regulators of macrophages. Prostaglandin E3 (PGE3) is an eicosanoid derived from eicosapentaenoic acid, which is converted by cyclooxygenase, followed by prostaglandin E synthase successively. We found that PGE3 played an anti‐inflammatory role by inhibiting LPS and interferon‐γ‐induced M1 polarization and promoting interleukin‐4‐mediated M2 polarization (M2a). Further, we found that although PGE3 had no direct effect on the growth of prostate cancer cells in vitro, PGE3 could inhibit prostate cancer in vivo in a nude mouse model of neoplasia. Notably, we found that PGE3 significantly inhibited prostate cancer cell growth in a cancer cell‐macrophage co‐culture system. Experimental results showed that PGE3 inhibited the polarization of tumour‐associated M2 macrophages (TAM), consequently producing indirect anti‐tumour activity. Mechanistically, we identified that PGE3 regulated the expression and activation of protein kinase A, which is critical for macrophage polarization. In summary, this study indicates that PGE3 can selectively promote M2a polarization, while inhibiting M1 and TAM polarization, thus exerting an anti‐inflammatory effect and anti‐tumour effect in prostate cancer.

Bladder cancer (BLCA) remains the leading cause of cancer‐related mortality among genitourinary malignancies worldwide. BLCA metastasis represents the primary reason for its poor prognosis. In this study, we report that decreased expression of partitioning defective 3 (Par3), a polarity protein (encoded by PARD3), is associated with tumor aggressive phenotypes and poor prognosis in BLCA patients. Consistently, ablation of Par3 promotes the metastasis and invasion of BLCA cells in vitro and in vivo. Further studies reveal that zinc finger protein Snail represses the expression of Par3 by binding to E2‐box (CAGGTG) of PARD3 promoter‐proximal. Inhibition of GSK‐3β promotes the expression and nuclear localization of Snail and then reduces the expression of Par3, resulting in the metastasis and invasion of BLCA cells. Moreover, we detected the interaction between Par3 (936‐1356 aa) and ZO‐1 (1372‐1748 aa), which is involved in the maintenance of tight junction. Together, our results demonstrate that the GSK‐3β/Snail/Par3/ZO‐1 axis regulates BLCA metastasis, and Snail is a major regulator for Par3 protein expression in BLCA., We report here that (a) polarity protein Par3 is frequently lost in BLCA patients and is associated with muscle‐invasive phenotypes and poor prognosis; (b) Par3 deficiency is required for BLCA invasion and metastasis in vitro and in vivo; (c) Snail suppresses Par3 expression through its binding to PARD3 E2‐box; and (d) overexpression of Par3 abolishes LiCl‐ or Snail‐induced BLCA metastatic behaviors, suggesting a role of the GSK‐3β/Snail/Par3 axis in regulating BLCA metastasis.

Distinct morphological subtypes of colorectal cancer (CRC) confer a bleak clinical outlook. In a recent issue of The Journal of Pathology, Onuma et al investigated morphological evolution of a highly fatal CRC subtype known as micropapillary cancer (MPC). This study enhances understanding of MPC biology including essential regulatory signals, cellular and multicellular phenotypes, as well as cancer behaviour. Iterative modelling in three-dimensional (3D) patient-derived CRC tissue-originated spheroids (CTOSs) revealed spatiotemporal oscillations of Rho-ROCK hyperactivity underlying reversal of membrane polarity and suppression of lumen formation during development of multicellular MPC morphology. Corroborative studies in CTOSs, xenografts, and archival human CRCs confirm human disease relevance. Although cancer morphology has previously been considered irreversible, targeted inhibition of Rho-ROCK activity restored membrane polarity, lumenized multicellular assembly, and suppressed MPC morphology in 3D CTOS cultures and xenografts. Collectively, the study identifies molecular, biophysical, and multicellular mechanisms implicated in morphological evolution of micropapillary CRC. © 2021 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Several signaling pathways were involved in M1 (classic) and M2 (alternative) macrophage polarization. Disruption of M2-related signaling pathways and improvement of M1-related signaling pathways can be identified as one of the cancer therapeutic approaches. Prevention of macrophage differentiation into M2 by different herbal agents with antitumor properties can be considered as a promising therapeutic target for cancer patients. In the present review study, we investigated the effect of herbal compounds on M1 and M2 related signaling pathways to reduce M2 and increase M1 macrophage polarization for the treatment of different types of cancer.

Cytochrome P450 2A6 (CYP2A6) is an important metabolic enzyme and is involved in the progression of hepatocellular carcinoma (HCC). However, its specific function and the mechanism of modulation remain to be elucidated. In this study, we found that CYP2A6 is dramatically downregulated in HCC. CYP2A6 expression is closely associated with pathological grading, histologic grade, hepatitis, vascular metastasis, liver inflammation, and worse prognosis. Reduced expression of CYP2A6 contributes to alternative activation of macrophage polarization and impairs macrophage maturation and phagocytosis. Mechanistically, CYP2A6 participates in arachidonic acid metabolism, initiates 20‐hydroxyeicosatetraenoic acid (HETE) generation, and inhibits epoxyeicosatrienoic acid (EET) generation. Disruption of the equilibrium between 20‐HETE and EETs can induce macrophage polarization, thereby modulating antitumor immunity., Cytochrome P450 2A6 (CYP2A6) has an antitumor role and is dramatically downregulated in HCC. CYP2A6 participates in arachidonic acid metabolism, initiates 20‐hydroxyeicosatetraenoic acid (HETE) generation, and inhibits epoxyeicosatrienoic acid (EET) generation. Disruption of the equilibrium between 20‐HETE and EETs can induce macrophage polarization, thereby modulating antitumor immunity.

Extracellular vesicles (EVs), a broad term for the lipid microparticles known as microvesicles and exosomes, are discharged by cells into their surrounding space. Microvesicles are discharged upon outward plasma membrane budding, while exosomes are secreted after multivesicular body (MVB) fusion with the plasma membrane. The majority of information regarding EV biology comes from studies performed in non-polarized cells. Here we characterize EV release in polarized cells. We found a substantial asymmetry in the number and composition of EVs produced and released from the apical membrane of epithelial cells as compared to the basolateral membrane. We showed that the quantitative difference is related to the polarized distribution of two phosphoinositide species between the two cell surfaces and that the peculiar biochemical composition of resultant EVs reflects their site of origin. In particular, apical and basolateral exosomes may derive from distinct classes of MVBs originating from and fusing with the same plasma membrane. We identify VAMP8/Endobrevin as a regulator of the basolateral release of exosomes, whereas the mechanism responsible for apical EV release requires further study.

Aims Gastric dysplasia is a risk factor for synchronous and subsequent gastric carcinoma. Distinguishing gastric dysplasia from reactive changes is subject to interobserver disagreement and is a frequent reason for expert consultation. We previously used assessment of surface cell polarity (the 'four lines') as a key feature to decrease equivocal diagnoses in Barrett oesophagus. In the current study, we examined for the presence or absence of the four lines in gastric dysplasia and reactive gastropathy. Materials and methods The study includes all (n = 91) in-house biopsies with at least gastric dysplasia from the surgical pathology archives of two academic institutions during a 5-year period from 2008 to 2012. A reactive gastropathy group (n = 60) was created for comparison. Results The dysplasia/neoplasia group was comprised of 14 biopsies of gastric foveolar-type dysplasia, 59 of intestinal-type dysplasia, 14 with dysplasia in fundic gland polyps, three pyloric gland adenomas and one oxyntic gland adenoma. Loss of surface cell polarity was seen in all 88 dysplasia cases with evaluable surface epithelium. All 57 reactive gastropathy cases with evaluable surface epithelium showed intact surface cell polarity except in focal areas directly adjacent to erosions in 17 cases, where the thin wisp of residual surface mucin could not be appreciated on haematoxylin and eosin. Conclusion Surface cell polarity (the four lines) was lost in all gastric dysplasia biopsies with evaluable surface epithelium and maintained in all biopsies of reactive gastropathy. Caution should be taken in using this feature adjacent to erosions in reactive gastropathy.

Type XVII collagen (COL17) is a transmembrane protein expressed in the basal epidermis. COL17 serves as a niche for epidermal stem cells, and although its reduction has been implicated in altering cell polarity and ageing of the epidermis, it is unknown how COL17 affects epidermal cell polarity. Here, we uncovered COL17 as a binding partner of the aPKC-PAR complex, which is a key regulating factor of cell polarity. Immunoprecipitation-immunoblot assay and protein-protein binding assay revealed that COL17 interacts with aPKC and PAR3. COL17 deficiency or epidermis-specific aPKCλ deletion destabilized PAR3 distribution in the epidermis, while aPKCζ knockout did not. Asymmetrical cell division was pronounced in COL17-null neonatal paw epidermis. These results show that COL17 is pivotal for maintaining epidermal cell polarity. Our study highlights the previously unrecognized role of COL17 in the basal keratinocytes.

Background Macrophage polarization affects tumor growth, metabolism, and many other tumor processes. M1 macrophages can promote antitumor immunity response. Signal transducer and activator of transcription 1 (STAT1) is one of the critical transcription factors in this process, which promotes the expression of a series of inflammatory molecules. STAT1 has been reported as a potential target of reactive oxygen species (ROS)‐induced glutathionylation, while the glutathionylation of STAT1 in macrophages and its underlying regulatory mechanism remains unclear. Glutaredoxin 1 (Grx1) functions as a deglutathionylation enzyme, which regulates the activities of many proteins through reversing glutathionylation. Methods GeneChip and RT‐qPCR was first applied to test the mRNA level of Grx1 in M1 macrophages. Western blot was then used to evaluate the variations of the Grx1 protein expression in M1 macrophages. Next, immunoprecipitation was used to investigate the glutathionylated STAT1 in both wild‐type and Grx1−/− mouse macrophages. Finally, the induced alterations of STAT1 activity and function by Grx1 in M1 macrophage were examined by western blot and RT‐qPCR. Results In M1‐type macrophages, the levels of Grx1 were elevated. Glutathionylation of STAT1 was negatively regulated by Grx1. Furthermore, depletion of Grx1 increased the activity of STAT1, and thereby promoted the mRNA level of C‐X‐C motif chemokine ligand 9 (CXCL9) during M1‐type polarization of macrophages. Conclusions Grx1 controlled deglutathionylation of STAT1, which in turn might regulate M1‐type polarization of macrophages., In M1‐type macrophages, the levels of Grx1 were elevated. Glutathionylation of STAT1 was negatively regulated by Grx1. Furthermore, depletion of Grx1 increased the activity of STAT1, and promoted the mRNA levels of CXCL9 in the process of macrophages polarized to M1 phenotype.

Oncogenic mutations often trigger antitumor cellular response such as induction of apoptosis or cellular senescence. Studies in the last decade have identified the presence of the third guardian against mutation‐induced tumorigenesis, namely “cell competition.” Cell competition is a context‐dependent cell elimination whereby cells with higher fitness eliminate neighboring cells with lower fitness by inducing cell death. While oncogene‐induced apoptosis or oncogene‐induced senescence acts as a cell‐autonomous tumor suppressor, cell competition protects the tissue from tumorigenesis via cell‐cell communication. For instance, in Drosophila epithelium, oncogenic cells with cell polarity mutations overproliferate and develop into tumors on their own but are eliminated from the tissue when surrounded by wild‐type cells. Genetic studies in flies have unraveled that such tumor‐suppressive cell competition is regulated by at least three mechanisms: direct cell‐cell interaction between polarity‐deficient cells and wild‐type cells, secreted factors from epithelial cells, and systemic factors from distant organs. Molecular manipulation of tumor‐suppressive cell competition could provide a novel therapeutic strategy against human cancers., Cell competition is a context‐dependent cell elimination whereby cells with higher fitness eliminate neighboring cells with lower fitness by inducing cell death. In Drosophila epithelium, oncogenic cells with cell polarity mutations overproliferate and develop into tumors on their own but are eliminated from the tissue when surrounded by wild‐type cells. Genetic studies in flies have unraveled that such tumor‐suppressive cell competition is regulated by at least three mechanisms: direct cell‐cell interaction between polarity‐deficient cells and wild‐type cells, secreted factors from epithelial cells, and systemic factors from distant organs.

The effects of mesenchymal stem cells (MSCs) on different types of diseases are controversial, and the inner mechanisms remain unknown, which retards the utilization of MSCs in disease therapy. In this study, we aimed to elucidate the mechanisms of MSCs‐extracellular vesicles (EVs) carrying transforming growth factor‐beta 1 (TGF‐β1) in M2 polarization in mouse macrophages via the microRNA‐132 (miR‐132)/E3 ubiquitin ligase myc binding protein 2 (Mycbp2)/tuberous sclerosis complex 2 (TSC2) axis. Mouse MSCs were isolated for adipogenic and osteogenic induction, followed by co‐culture with mouse macrophages RAW264.7. Besides, mouse macrophages RAW264.7 were co‐cultured with MSCs‐EVs in vitro, where the proportion of macrophages and inflammation were detected by flow cytometry and ELISA. The experimental data revealed that MSCs‐EVs promoted M2 polarization of macrophages, and elevated interleukin (IL)‐10 expression and inhibited levels of IL‐1β, tumour necrosis factor (TNF)‐α and IL‐6. MSC‐EV‐treated macrophages RAW264.7 increased TGF‐β1 expression, thus elevating miR‐132 expression. MiR‐132 directly bound to Mycbp2, as confirmed by luciferase activity assay. Meanwhile, E3 ubiquitin ligase Mycbp2 could ubiquitinate TSC2 protein. Furthermore, silencing TGF‐β1 inhibited M2 polarization of MSC‐EV‐treated macrophages. Taken conjointly, this study provides evidence reporting that MSC‐secreted EVs carry TGF‐β1 to promote M2 polarization of macrophages via modulation of the miR‐132/Mycbp2/TSC2 axis.

Objective After bone prosthesis replacement, M1-type macrophage polarization can be induced by titanium (Ti) particles and produce inflammatory, leading to osteolysis. Adipocyte-derived exosomes (ADEs) exert immune-modulatory impact on the macrophage, while whether it can inhibit the macrophage polarization induced by Ti is unclear. This study focuses on the M1-type macrophage and aims to determine the effect of ADEs on Ti-induced M1-type macrophage polarization in osteolytic mice and the involved mechanism. Methods Ti particle-induced osteolysis mouse model was established and macrophages were isolated from the osteolysis site. The levels of NLRP3 and specific markers for M1-type macrophage were determined. ADEs isolated from adipocyte cell line 3T3-L1, or conditioned ADEs with low-expressed miR-34a isolated from 3T3-L1 transfected with miR-34a inhibitor were co-cultured with RAW 264.7 to determine their impact on the polarization of macrophage. Results ADEs reduced the M1-type macrophage polarization and caused the upregulation of miR-34a in macrophage of the osteolysis site of the osteolysis mouse model. Also, the level of miR-34a in ADEs was higher than that in the adipocyte. The conditioned ADEs expressed a low level of miR-34a and boosted the Ti-induced M1-type polarization. MiR-34a could target NLRP3 and negatively regulated its expression. Moreover, NLRP3 knockdown in macrophage restricted the conditioned ADEs to promote macrophage towards to Ti-induced M1-type polarization. The inhibitory function of ADEs on M1-type macrophage polarization was abolished by miR-34a silencing in the mouse osteolysis model. Conclusion The miR-34a carried by ADEs reduced the polarization of M1-type macrophages by targeting macrophage NLRP3 during Ti particle-induced osteolysis.

The progression of lung cancer is majorly facilitated by TAMs (tumour‐associated macrophages). However, how the TAMs infiltrate the NSCLC microenvironment and the associated biochemical are not fully elaborated. Research has revealed that changes in CtBP1 modulates innate immunity. Here, we investigated if CtBP1 facilitates infiltration of TAM and the subsequent progression of NSCLC. Immunohistochemical analysis was carried out in 96 NSCLC patients to estimate the clinicopathological importance of CtBP1 in the disease. CtBP1 overexpression and knockdown were carried out to assess the activity of CtBP1 in NSCLC cells. Elevated expression of CtBP1 correlated positively with TAMs infiltration into NSCLC tissues, induced EMT (epithelial‐mesenchymal transition) in NSCLC cells and modulated the activated NF‐κB signalling pathway leading to increase in CCL2 secretion from NSCLC cells, thus promoting TAM recruitment and polarization. TAM induction and polarization reduced significantly on exhausting p65 in NSCLC cells with CtBP1. Moreover, infiltration of TMAs was reduced remarkably on antagonist‐mediated blocking of CCR2 and impeded the progression of NSCLC in a mouse model. These findings thus show a novel insight into the process of CtBP1‐regulated TAM infiltration in NSCLC.

Macrophages are key regulators of the development and progression of asthma, facilitating deleterious airway remodeling in affected patients. Immune cell function is tightly regulated by microRNAs (miRNAs), but how these miRNAs impact macrophage-mediated airway remodeling in the context of asthma remains to be determined. In this study, we utilized an ovalbumin (OVA)-based murine model of asthma to evaluate the importance of miRNAs within these macrophages. We found that macrophages from mice that had been sensitized with and exposed to OVA expressed higher levels of M2-like phenotypic markers and exhibited significantly altered expression of both miR-142-5p and miR-130a-3p. When these isolated pulmonary macrophages were cultured in vitro, we determined that transfecting them with miR-142-5p antisense oligonucleotide (ASO) or miR-130a-3p mimics was sufficient to inhibit the ability of interleukin-4 to induce M2 cytokine production. We additionally confirmed the in vivo relevance of these miRNAs in a Ccr2-/- murine model system mimicking asthma. Specifically, we determined that transfecting monocytes with miR-142-5p ASO and/or miR-130a-3p mimics was sufficient to disrupt the ability of these cells to promote airway remodeling. As such, these findings reveal that miR-142-5p and miR-130a-3p dysregulation are important factors governing the polarization of macrophages and associated airway remodeling in OVA-sensitized mice.

Two small Ras-like GTPases, MglA and SofG, work in synchrony to drive cell polarity and motility in the soil bacterium, Myxococcus xanthus. While MglA regulates two types of motility in Myxococcus and drives cell polarity reversals, SofG regulates social motility enabled by the type IV pili (T4P) machinery. In order to understand the molecular basis of how multiple GTPases act concertedly, we initiated biochemical studies on SofG. A construct of SofG (SofG∆60 ) was purified as a homogenous monomer and could bind to GDP and GTP. Intrinsic GTP hydrolysis by SofG∆60 was negligible. Earlier work from the laboratory revealed that MglB functions both as a GTPase-activating protein (GAP) and a guanine nucleotide exchange factor (GEF) for MglA. Biochemical assays of SofG∆60 established that MglB interacts with GTP-bound SofG∆60 and acts as a GAP for SofG∆60 . Interaction of MglB with SofG∆60 in the GDP-bound conformation was not observed, thereby suggesting that MglB might not act as a GEF for SofG∆60 . The existence of a common GAP for both SofG and MglA could potentially contribute to concerted regulation of their GTPase activities, and mediate crosstalk between the two GTPases involved in motility of M. xanthus. Sequence analysis revealed the features for a SofG-like subclass of prokaryotic small Ras-like GTPases that enable MglB to act as a dual-specificity GAP.

Acute respiratory distress syndrome (ARDS) is a fatal disease characterized by excessive infiltration of inflammatory cells. MCTR1 is an endogenously pro‐resolution lipid mediator. We tested the hypothesis that MCTR1 accelerates inflammation resolution through resident M2 alveolar macrophage polarization. The mice received MCTR1 via intraperitoneal administration 3 days after LPS stimulation, and then, the bronchoalveolar lavage (BAL) fluid was collected 24 hours later to measure the neutrophil numbers. Flow cytometry was used to sort the resident and recruited macrophages. Post‐treatment with MCTR1 offered dramatic benefits in the resolution phase of LPS‐induced lung injury, including decreased neutrophil numbers, reduced BAL fluid protein and albumin concentrations and reduced histological injury. In addition, the expression of the M2 markers Arg1, FIZZ1, Remlα, CD206 and Dectin‐1 was increased on resident macrophages in the LPS + MCTR1 group. Resident macrophage depletion abrogated the therapeutic effects of MCTR1, and reinjection of the sorted resident macrophages into the lung decreased neutrophil numbers. Finally, treatment with MCTR1 increased STAT6 phosphorylation. The STAT6 inhibitor AS1517499 abolished the beneficial effects of MCTR1. In conclusion, MCTR1 promotes resident M2 alveolar macrophage polarization via the STAT6 pathway to accelerate resolution of LPS‐induced lung injury.

It has been documented that M2 macrophage polarization plays a suppressive role in atherosclerosis in diabetes mellitus (DM). In addition, prostaglandin E2 (PGE2) is implicated in the development of M2 macrophage polarization. Therefore, the study aimed to investigate the specific mechanism of PGE2 in M2 macrophage polarization in diabetic coronary atherosclerosis (DMAS). Initially, clinical samples were obtained and DMAS mouse model was established. The expression of BDNF was determined, and M1 and M2 macrophage polarizations were evaluated. Then, the levels of BDNF and PGE2 were modified in DMAS mice and the serum indicator, atherosclerotic plaque, lipid uptake by PBMCs, as well as M1 and M2 macrophage polarization were determined. Macrophages were isolated and the effects of PGE2 and the CREB/BDNF/TrkB signaling pathway on M2 macrophage polarization were explored. BDNF was downregulated and macrophages were differentiated into M1 in DMAS patients and mice. BDNF and PGE2 were observed to promote M2 macrophage polarization, where atherosclerotic plaque and lipid uptake by PBMCs were reduced, and DMAS was alleviated in mice. Overexpression of BDNF activated the CREB/BDNF/TrkB signaling pathway and stimulated M2 macrophage polarization in macrophages. PGE2 stimulated M2 macrophage polarization by inducing KLF4 via the activation of the CREB/BDNF/TrkB signaling pathway. This study demonstrates that PGE2 promotes M2 macrophage polarization by activating the CREB/BDNF/TrkB signaling pathway, thus alleviating DMAS.

Colorectal cancer (CRC) is the third most common solid tumor worldwide and has shown resistance to several immunotherapies, particularly immune checkpoint blockade therapy, which is effective in many other types of cancer. Our previous studies indicated that the active fraction of Garcinia yunnanensis (YTE-17), had potent anticancer activities by regulating multiple signaling pathways. However, knowledge regarding the mechanism and effect of YTE-17 in the prevention of CRC is limited. This study tested the effects of YTE-17 on colon cancer development in vivo by using two murine models: the carcigenic azoxymethane/dextran sulfate sodium (AOM/DSS)-induced CRC model and a genetically induced model using ApcMin/+ mice. Here, the tumor load, tumor number, histology, and even some oncogenes were used to evaluate the effect of YTE-17. The intragastric administration of YTE-17 for 12 weeks significantly decreased CRC incidence, tumor number and size, immunity, and some tumor-associated macrophage (TAM) markers, including CD206, Arg-1, IL-10, and TGF-β. Importantly, the macrophages depletion by clodronate (CEL) also played a role in reducing the tumor burden and inhibiting tumor development, which were not affected by YTE-17 in the ApcMin/+ mice. Moreover, the YTE-17 treatment attenuated CRC cell growth in a co-culture system in the presence of macrophages. Consistently, YTE-17 effectively reduced the tumor burden and macrophage infiltration and enhanced immunity in the AOM/DSS and ApcMin/+ colon tumor models. Altogether, we demonstrate that macrophages in the microenvironment may contribute to the development and progression of CRC cells and propose YTE-17 as a new potential drug option for the treatment of CRC.

Hepatocytes, the main epithelial cells of the liver, organize their polarized membrane domains differently from ductal epithelia. They also differ in their biosynthetic delivery of single-membrane-spanning and glycophosphatidylinositol-anchored proteins to the apical domain. While ductal epithelia target apical proteins to varying degrees from the trans-Golgi network (TGN) to the apical surface directly, hepatocytes target them first to the basolateral domain, from where they undergo basolateral-to-apical transcytosis. How TGN-to-surface transport differs in both scenarios is unknown. Here, we report that the basolateral detour of a hepatocyte apical protein is due, in part, to low RhoA activity at the TGN, which prevents its segregation from basolateral transport carriers. Activating Rho in hepatocytic cells, which switches their polarity from hepatocytic to ductal, also led to apical-basolateral cargo segregation at the TGN as is typical for ductal cells, affirming a central role for Rho-signaling in different aspects of the hepatocytic polarity phenotype. Nevertheless, Rho-induced cargo segregation was not sufficient to target the apical protein directly; thus, failure to recruit apical targeting machinery also contributes to its indirect itinerary.

MicroRNAs (miRNAs) provide context-dependent transcriptional regulation of genes comprising signalling networks throughout the developing organism including morphogenesis of the embryonic neural tube (NT). Using a high-sensitivity, high-coverage microarray analysis platform, miRNA expression in the murine embryonic NT during the critical stages of its formation was examined. Analysis of a number of differentially expressed (DE) miRNAs enabled identification of several gene targets associated with cellular processes essential for normal NT development. Using computational pathway analysis, interactive biologic networks and functional relationships connecting DE miRNAs with their targeted messenger RNAs (mRNAs) were identified. Potential mRNA targets and a key signal transduction pathway governing critical cellular processes indispensable for normal mammalian neurulation were also identified. RNA preparations were also used to hybridize both miRNA arrays and mRNA arrays allowing miRNA-mRNA target analysis using data of DE miRNAs and DE mRNAs - co-expressed in the same developing NT tissue samples. Identification of these miRNA targets provides key insight into the epigenetic regulation of NT development as well as into potential mechanistic underpinning of NT defects. SIGNIFICANCE OF THE STUDY: This study underscores the premise that microRNAs are potential coordinators of normal neural tube (NT) formation, via regulation of the crucial, planar cell polarity pathway. Any alteration in their expression during neurulation would result in abnormal NT development.

Defective Kernel 1 (DEK1) is genetically at the nexus of the 3D morphogenesis of land plants. We aimed to localize DEK1 in the moss Physcomitrella patens to decipher its function during this process. To detect DEK1 in vivo, we inserted the tdTomato fluorophore into PpDEK1 gene locus. Confocal microscopy coupled with the use of time-gating allowed the precise DEK1 subcellular localization during 3D morphogenesis. DEK1 localization displays a strong polarized signal, as it is restricted to the plasma membrane domain between recently divided cells during the early steps of 3D growth development as well as during the subsequent vegetative growth. The signal furthermore displays a clear developmental pattern because it is only detectable in recently divided and elongating cells. Additionally, DEK1 localization appears to be independent of its calpain domain proteolytic activity. The DEK1 polar subcellular distribution in 3D tissue developing cells defines a functional cellular framework to explain its role in this developmental phase. Also, the observation of DEK1 during spermatogenesis suggests another biological function for this protein in plants. Finally the DEK1-tagged strain generated here provides a biological platform upon which further investigations into 3D developmental processes can be performed.

Aseptic loosening caused by wear particles is a common complication after total hip arthroplasty. We investigated the effect of the quercetin on wear particle‐mediated macrophage polarization, inflammatory response and osteolysis. In vitro, we verified that Ti particles promoted the differentiation of RAW264.7 cells into M1 macrophages through p‐38α/β signalling pathway by using flow cytometry, immunofluorescence assay and small interfering p‐38α/β RNA. We used enzyme‐linked immunosorbent assays to confirm that the protein expression of M1 macrophages increased in the presence of Ti particles and that these pro‐inflammatory factors further regulated the imbalance of OPG/RANKL and promoted the differentiation of osteoclasts. However, this could be suppressed, and the protein expression of M2 macrophages was increased by the presence of the quercetin. In vivo, we revealed similar results in the mouse skull by μ‐CT, H&E staining, immunohistochemistry and immunofluorescence assay. We obtained samples from patients with osteolytic tissue. Immunofluorescence analysis indicated that most of the macrophages surrounding the wear particles were M1 macrophages and that pro‐inflammatory factors were released. Titanium particle‐mediated M1 macrophage polarization, which caused the release of pro‐inflammatory factors through the p‐38α/β signalling pathway, regulated OPG/RANKL balance. Macrophage polarization is expected to become a new clinical drug therapeutic target.

Resident alveolar macrophages (AMs) suppress allergic inflammation in murine asthma models. Previously we reported that resident AMs can blunt inflammatory signaling in alveolar epithelial cells (ECs) by transcellular delivery of suppressor of cytokine signaling 3 (SOCS3) within extracellular vesicles (EVs). Here we examined the role of vesicular SOCS3 secretion as a mechanism by which AMs restrain allergic inflammatory responses in airway ECs. Bronchoalveolar lavage fluid (BALF) levels of SOCS3 were reduced in asthmatics and in allergen-challenged mice. Ex vivo SOCS3 secretion was reduced in AMs from challenged mice and this defect was mimicked by exposing normal AMs to cytokines associated with allergic inflammation. Both AM-derived EVs and synthetic SOCS3 liposomes inhibited the activation of STAT3 and STAT6 as well as cytokine gene expression in ECs challenged with IL-4/IL-13 and house dust mite (HDM) extract. This suppressive effect of EVs was lost when they were obtained from AMs exposed to allergic inflammation-associated cytokines. Finally, inflammatory cell recruitment and cytokine generation in the lungs of OVA-challenged mice were attenuated by intrapulmonary pretreatment with SOCS3 liposomes. Overall, AM secretion of SOCS3 within EVs serves as a brake on airway EC responses during allergic inflammation, but is impaired in asthma. Synthetic liposomes encapsulating SOCS3 can rescue this defect and may serve as a framework for novel therapeutic approaches targeting airway inflammation.

Retention after treatment and effective anchorage control are two essential factors in orthodontics. Our study aimed to explore the effects of fucoidan on orthodontic tooth movement (OTM) and the involvement of macrophages. We established a murine OTM model to test the effect of fucoidan administration. We found that mice injected with fucoidan had a deceleration in OTM and a higher bone mineral density. Moreover, fucoidan increased the proportion of F4/80+ CD206+ macrophages and promoted the messenger RNA expression of Arg-1, CD206, and IL-10 at both in vivo and in vitro levels. In addition, macrophages showed lower expression of TNF-α, IL-1β, and IL-6 and a decrease in F4/80+ CD11c+ cells. Mechanistically, the level of phosphorylated STAT3 was elevated in unpolarized and restorative macrophages after treatment with fucoidan. Taken together, our findings suggest that fucoidan treatment inhibits OTM and enhances the stability of teeth after movement by promoting restorative macrophages through the STAT3 pathway.

Reperfusion of the ischemic intestine often leads to drive distant organ injury, especially injuries associated with hepatocellular dysfunction. The precise molecular mechanisms and effective multiple organ protection strategies remain to be developed. In the current study, significant remote liver dysfunction was found after 6 hours of reperfusion according to increased histopathological scores, serum lactate dehydrogenase (LDH), alanine aminotransferase (ALT)/aspartate aminotransferase (AST) levels, as well as enhanced bacterial translocation in a rat intestinal ischemia/reperfusion (I/R) injury model. Moreover, receptor-interacting protein kinase 1/3 (RIP1/3) and phosphorylated-MLKL expressions in tissue were greatly elevated, indicating that necroptosis occurred and resulted in acute remote liver function impairment. Inhibiting the necroptotic pathway attenuated HMGB1 cytoplasm translocation and tissue damage. Meanwhile, macrophage-depletion study demonstrated that Kupffer cells (KCs) are responsible for liver damage. Blocking HMGB1 partially restored the liver function via suppressed hepatocyte necroptosis, tissue inflammation, hepatic KCs, and circulating macrophages M1 polarization. What's more, HMGB1 neutralization further protects against intestinal I/R-associated liver damage in microbiota-depleted rats. Therefore, intestinal I/R is likely associated with acute liver damage due to hepatocyte necroptosis, and which could be ameliorated by Nec-1 administration and HMGB1 inhibition with the neutralizing antibody and inhibitor. Necroptosis inhibition and HMGB1 neutralization/inhibition, may emerge as effective pharmacological therapies to minimize intestinal I/R-induced acute remote organ dysfunction.

Owing to the complexity of the pathophysiological mechanisms driving epileptogenesis following traumatic brain injury (TBI), effective preventive treatment approaches are not yet available for posttraumatic epilepsy (PTE). Neuroinflammation appears to play a critical role in the pathogenesis of the acquired epilepsies, including PTE, but despite a large preclinical literature demonstrating the ability of anti-inflammatory treatments to suppress epileptogenesis and chronic seizures, no anti-inflammatory treatment approaches have been clinically proven to date. TBI triggers robust inflammatory cascades, suggesting that they may be relevant for the pathogenesis of PTE. A major cell type involved in such cascades is the microglial cells-brain-resident immune cells that become activated after brain injury. When activated, these cells can oscillate between different phenotypes, and such polarization states are associated with the release of various pro- and anti-inflammatory mediators that may influence brain repair processes, and also differentially contribute to the development of PTE. As the molecular mechanisms and key signaling molecules associated with microglial polarization in brain are discovered, strategies are now emerging that can modulate this polarization, promoting this as a potential therapeutic strategy for PTE. In this review, we discuss the relevant literature regarding the polarization of brain-resident immune cells following TBI and attempt to put into perspective a role in epilepsy pathogenesis. Finally, we explore potential strategies that could polarize microglia/macrophages toward a neuroprotective phenotype to mitigate PTE development.

Background Fibroblasts executing directional migration position their centrosome, and their Golgi apparatus, in front of the nucleus towards the cell leading edge. Centrosome positioning relative to the nucleus has been associated to mechanical forces exerted on the centrosome by the microtubule-dependent molecular motor cytoplasmic dynein 1, and to nuclear movements such as rearward displacement and rotation events. Dynein has been proposed to regulate the position of the centrosome by exerting pulling forces on microtubules from the cell leading edge, where the motor is enriched during migration. However, the mechanism explaining how dynein acts at the front of the cells has not been elucidated. Results We present here results showing that the protein Focal Adhesion Kinase (FAK) interacts with dynein and regulates the enrichment of the dynein/dynactin complex at focal adhesions at the cell the leading edge of migrating fibroblasts. This suggests that focal adhesions provide anchoring sites for dynein during the polarisation process. In support of this, we present evidence indicating that the interaction between FAK and dynein, which is regulated by the phosphorylation of FAK on its Ser732 residue, is required for proper centrosome positioning. Our results further show that the polarisation of the centrosome can occur independently of nuclear movements. Although FAK regulates both nuclear and centrosome motilities, downregulating the interaction between FAK and dynein affects only the nuclear independent polarisation of the centrosome. Conclusions Our work highlights the role of FAK as a key player in the regulation of several aspects of cell polarity. We thus propose a model in which the transient localisation of dynein with focal adhesions provides a tuneable mechanism to bias dynein traction forces on microtubules allowing proper centrosome positioning in front of the nucleus. Significance We unravel here a new role for the cancer therapeutic target FAK in the regulation of cell morphogenesis.

Wear particle‐stimulated inflammatory bone destruction and the consequent aseptic loosening remain the primary causes of artificial prosthesis failure and revision. Previous studies have demonstrated that curcumin has a protective effect on bone disorders and inflammatory diseases and can ameliorate polymethylmethacrylate‐induced osteolysis in vivo. However, the effect on immunomodulation and the definitive mechanism by which curcumin reduces the receptor activators of nuclear factor‐kappa B ligand (RANKL)‐stimulated osteoclast formation and prevents the activation of osteoclastic signalling pathways are unclear. In this work, the immunomodulation effect and anti‐osteoclastogenesis capacities exerted by curcumin on titanium nanoparticle‐stimulated macrophage polarization and on RANKL‐mediated osteoclast activation and differentiation in osteoclastic precursor cells in vitro were investigated. As expected, curcumin inhibited RANKL‐stimulated osteoclast maturation and formation and had an immunomodulatory effect on macrophage polarization in vitro. Furthermore, studies aimed to identify the potential molecular and cellular mechanisms revealed that this protective effect of curcumin on osteoclastogenesis occurred through the amelioration of the activation of Akt/NF‐κB/NFATc1 pathways. Additionally, an in vivo mouse calvarial bone destruction model further confirmed that curcumin ameliorated the severity of titanium nanoparticle‐stimulated bone loss and destruction. Our results conclusively indicated that curcumin, a major biologic component of Curcuma longa with anti‐inflammatory and immunomodulatory properties, may serve as a potential therapeutic agent for osteoclastic diseases.

The efficacy of cardiac regenerative strategies for myocardial infarction (MI) treatment is greatly limited by the cardiac microenvironment. The combination of reactive oxygen species (ROS) scavenging to suppress the oxidative stress damage and macrophage polarization to regenerative M2 phenotype in the MI microenvironment can be desirable for MI treatment. Herein, melanin nanoparticles (MNPs)/alginate (Alg) hydrogels composed of two marine‐derived natural biomaterials, MNPs obtained from cuttlefish ink and alginate extracted from ocean algae, are proposed. Taking advantage of the antioxidant property of MNPs and mechanical support from injectable alginate hydrogels, the MNPs/Alg hydrogel is explored for cardiac repair by regulating the MI microenvironment. The MNPs/Alg hydrogel is found to eliminate ROS against oxidative stress injury of cardiomyocytes. More interestingly, the macrophage polarization to regenerative M2 macrophages can be greatly promoted in the presence of MNPs/Alg hydrogel. An MI rat model is utilized to evaluate the feasibility of the as‐prepared MNPs/Alg hydrogel for cardiac repair in vivo. The antioxidant, anti‐inflammatory, and proangiogenesis effects of the hydrogel are investigated in detail. The present study opens up a new way to utilize natural biomaterials for MI treatment and allows to rerecognize the great value of natural biomaterials in cardiac repair., Natural melanin nanoparticles/alginate hydrogels are proposed to attenuate inflammation by scavenging harmful reactive oxygen species and promoting macrophage polarization to regenerative M2 phenotype in the MI region for cardiac repair.

Glioma is regarded as an aggressive lethal primary brain tumor. Jumonji domain containing 1C (JMJD1C) is a H3K9 demethylase which participates in the progression of various tumors, but its specific function and underlying mechanism in glioma development remain undefined, which is the purpose of our work. We initially assessed JMJD1C expression in glioma tissues and cells using the assays of RT‐qPCR and immunohistochemistry. Meanwhile, the H3K9 level at the microRNA (miR)‐302a promoter region was measured by chromatin immunoprecipitation assay, while luciferase‐based reporter assay was performed for validation of the binding affinity between miR‐302a and methyltransferase‐like 3 (METTL3). The effect of METTL3 on suppressor of cytokine signaling 2 (SOCS2) was subsequently analyzed by MeRIP‐RT‐qPCR. Finally, a xenograft tumor model was established in nude mice, followed by measurement of tumor‐associated macrophages using flow cytometry. JMJD1C was poorly expressed in glioma tissues. Furthermore, JMJD1C increased miR‐302a expression through promoting H3K9me1 demethylation at the miR‐302a promoter region. miR‐302a was identified to target METTL3, which could inhibit SOCS2 expression via m6A modification. JMJD1C promoted M1 macrophage polarization and suppressed the growth of glioma xenografts through the miR‐302a/METTL3/SOCS2 axis both in vivo and in vitro. In conclusion, JMJD1C could enhance M1 macrophage polarization to inhibit the onset of glioma, bringing a new insight into the contribution of JMJD1C to the pathobiology of glioma, with possible implications for targeted therapeutic method., Our results indicate that JMJD1C enhances M1 macrophage polarization to inhibit the onset of glioma, which brings a new insight for the contributions of JMJD1C to the pathobiology of glioma.

ObjectivesMechanisms that govern angiogenesis are paramount to our understanding of how blood vessels are formed embryonically, maintained in adulthood and manifest disease. Akin to transcriptional regulation of endothelial-specific genes, vesicular trafficking events dictate protein localization, functional activity, and half-life, providing a critically important regulatory step. However, there is little information detailing endothelial-specific trafficking signatures. This is due, in part, by limitations in visualizing trafficking events in endothelial tissues. Our aim in this investigation was to explore the use of a 3-dimensional (3D) in vitro sprouting model to image and evaluate membrane trafficking events compared to the conventional 2-dimensional (2D)-based culture method.MethodsPrimary Human endothelial cells were challenged to make multicellular sprouts using a fibrin-bead assay. An assortment of cell polarity and Rab proteins were quantified via immunocytochemistry and live-imaging to compare their localization between 3D sprouts and 2D culture.ResultsOur results show that sprouts generated from the fibrin-bead assay grow close to the imaging plane allowing for an orthogonal view of apical and basal membrane domains. Compared with 2D culture in which the apical and basal domains are in the axial orientation, limiting resolution, 3D sprouts are acquired in the X-Y plane providing high-resolution for viewing trafficking events. Second, we demonstrate that fibrin-bead generated sprouts have a strong apicobasal polarity axis. Third, we directly compare imaging of trafficking mediators podocalyxin and Rab35 between 3D sprouts and 2D culture. Here, we show that 3D sprouting structures are well-suited to capture trafficking events that are not present in 2D culture due to the lack of a defined apical domain. Lastly, we compared exocytic events of von Willebrand Factor between 3D sprouting and 2D culture. Our results demonstrate a distinct imaging advantage for monitoring these trafficking programs in 3D sprouts as compared with conventional 2D culture.ConclusionsIn general, our results establish that the fibrin-bead sprouting assay is well-suited for sub-cellular imaging of trafficking events during angiogenic growth. Additionally, the 2D endothelial culture does not enforce the formation of an apicobasal polarity axis.

The cystic fibrosis transmembrane conductance regulator (CFTR) is an anion channel whose dysfunction causes cystic fibrosis (CF). The loss of CFTR function in pulmonary epithelial cells causes surface dehydration, mucus build‐up, inflammation, and bacterial infections that lead to lung failure. Little has been done to evaluate the effects of lipid perturbation on CFTR activity, despite CFTR residing in the plasma membrane. This work focuses on the acute effects of sphingomyelinase (SMase), a bacterial virulence factor secreted by CF relevant airway bacteria which degrades sphingomyelin into ceramide and phosphocholine, on the electrical circuitry of pulmonary epithelial monolayers. We report that basolateral SMase decreases CFTR‐mediated transepithelial anion secretion in both primary bronchial and tracheal epithelial cells from explant tissue, with current CFTR modulators unable to rescue this effect. Focusing on primary cells, we took a holistic ion homeostasis approach to determine a cause for reduced anion secretion following SMase treatment. Using impedance analysis, we determined that basolateral SMase inhibits apical and basolateral conductance in non‐CF primary cells without affecting paracellular permeability. In CF primary airway cells, correction with clinically relevant CFTR modulators did not prevent SMase‐mediated inhibition of CFTR currents. Furthermore, SMase was found to inhibit only apical conductance in these cells. Future work should determine the mechanism for SMase‐mediated inhibition of CFTR currents, and further explore the clinical relevance of SMase and sphingolipid imbalances., The loss of function of cystic fibrosis transmembrane conductance regulator (CFTR), in people with cystic fibrosis (CF), causes airway surface dehydration, mucus build‐up, inflammation, and bacterial infections that lead to lung failure. This work evaluates the effects of lipid perturbation on CFTR activity, focusing on the acute effects of sphingomyelinase (SMase), a bacterial virulence factor secreted by CF‐relevant airway bacteria, on the electrical circuitry of pulmonary epithelial monolayers. We report that basolateral SMase decreases CFTR‐mediated transepithelial anion secretion in both primary bronchial and tracheal epithelial cells from explant tissue, with current CFTR modulators unable to rescue this effect.

Allogeneic haematopoietic stem cell transplantation (allo‐HSCT) is the only curative method in treating haematologic malignant diseases. Graft‐versus‐host disease (GVHD) is a common complication post–allo‐HSCT, which can be life‐threatening. Mesenchymal stem cells (MSCs) as an adult stem cell with immunoregulatory function have demonstrated efficacy in steroid resistant acute GVHD (aGVHD). However, the outcome of aGVHD treated with MSCs in clinical trials varied and its underlying mechanism is still unclear. TGF‐β1 is a potent cytokine, which plays a key role in immunoregulation. In the present study, we firstly transduced the lentivirus vector containing TGF‐β1 gene with mouse bone marrow‐derived MSCs. Then, we investigated the immunosuppressive effect of TGF‐β1 gene‐modified MSCs on lymphocytes in vitro and its preventive and therapeutical effects on murine aGVHD model in vivo. Murine MSC was successfully isolated and identified. TGF‐β1 was efficiently transduced into mouse MSCs, and high level TGF‐β1 was detected. MSC‐TGF‐β1 shared the same morphology and immunotypic features of normal MSC. In vitro, MSC‐TGF‐β1 showed enhanced immunosuppressive function on lymphocyte proliferation. In vivo, MSC‐TGF‐β1 showed enhanced amelioration on the severity of aGVHD both in prophylactic and therapeutic murine models. Finally, the macrophages (MØs) derived from MSC‐TGF‐β1–treated mice showed a remarkably increasing of anti‐inflammatory M2‐like phenotype. Furthermore, the differentiation of CD4+ CD25+ Foxp3+ Treg cells was significantly increased in MSC‐TGF‐β1–treated group. Taken together, we proved that MSC‐TGF‐β1 showed enhanced alleviation of aGVHD severity in mice by skewing macrophages into a M2 like phenotype or increasing the proportion of Treg cells, which opens a new frontier in the treatment of aGVHD.

Perioperative neurocognitive disorders have been widely recognized as common adverse events after surgical intervention. Aging is one of the most important independent risk factors for worsened cognitive outcome, and this deterioration is linked to exacerbated microglia‐mediated neuroinflammation in the aged brain. Under pathological stimulation, microglia are capable of polarizing toward proinflammatory M1 and anti‐inflammatory M2 phenotypes. In the present study, we examined how aging affects microglial responses and neuroinflammation following peripheral surgery. Adult (2‐3 months) and aged (18 months old) male C57/BL6 mice were subjected to tibial fracture or sham surgery. Aged mice exhibited higher level of tumor necrosis factor‐α (TNF‐α) and interleukin‐1β (IL‐1β) in the hippocampus. The expression of synaptic protein synaptophysin (SYP) was also markedly reduced in the aged brain after the surgery. Both adult and aged mice showed significant increases in M1 microglial polarization (CD16/32). In contrast, tibial fracture surgery induced a decreased M2 microglial polarization (CD206, Ym1/2, Arg1) in aged brain but enhanced M2 microglial polarization in adult brain. Aged mice have upregulated voltage‐gated proton channel (Hv1) and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase subunit expression compared with adult mice. The percentage of CD16/32‐positive M1 microglia colabeling with Hv1 was higher in aged mice after tibial fracture surgery. Thus, Hv1/NADPH oxidase upregulation in the aged brain may shift the dynamic equilibrium of microglial activation toward M1 polarization and exaggerate postoperative neuroinflammatory responses after peripheral surgical intervention.

Breast cancer remains a leading cause of cancer-related death for women. The stepwise development of breast cancer through preinvasive to invasive disease is associated with progressive disruption of cellular and tissue organization. Apical-basal polarity is thought to be a barrier to breast cancer development, but the extent and potential mechanisms that contribute to disrupted polarity are incompletely understood. To investigate the cell polarity status of invasive breast cancers, we performed multiplex imaging of polarity markers on tissue cores from 432 patients from a spectrum of grades, stages and molecular subtypes. Apical-basal cell polarity was lost in 100% of cells in all cases studied, indicating that loss of epithelial polarity may be a universal feature of invasive breast cancer. We then analyzed genomic events from the TCGA dataset for an 18-gene set of core polarity genes. Coamplification of polarity genes with established breast oncogenes was found, which is consistent with functional cooperation within signaling amplicons. Gene-expression levels of several polarity genes were significantly associated with survival, and protein localization of Par6 correlated with higher grade, nodal metastasis and molecular subtype. Finally, multiple hotspot mutations in protein-protein interaction domains critical for cell polarity were identified. Our data indicate that genomic events likely contribute to pervasive disruption of epithelial polarity observed in invasive breast cancer.