Clinical Characteristics and Drug Resistance Mechanisms of Linezolid-Non-Susceptible Enterococcus in a Tertiary Hospital in Northwest China.

Purpose: To understand the detection rate and distribution characteristics of Linezolid-nonsusceptible Enterococcus (LNSE) and analyze the molecular typing and main drug resistance mechanisms of LNSE, providing a theoretical basis for the precision prevention and control of LNSE hospital infections. Methods: A total of 40 LNSE strains isolated from clinical specimens between January 1, 2012, and December 31, 2022, were collected. The LNSE isolates identified by instrument detection were confirmed using a microbroth dilution method. The WHONET 5.0 software was used for statistical analysis of LNSE detection rate, and the LNSE judgment was based on the 2022 CLSI criteria. PCR methods were used to detect 23S rRNA, cfr, optrA, and L3, L4 ribosomal RNA sites for linezolid resistance genes, and gene sequencing was used to verify the amplified PCR products. Multiple locus sequence typing (MLST) was performed to analyze the homology of LNSE strains. Results: A total of 6924 Enterococcus isolates were separated and identified from January 1, 2012, to December 31, 2022, of which 40 were LNSE strains (26 Enterococcus faecalis, 14 Enterococcus faecium), with a detection rate of 0.58% (40/6924). Among them, 28 Linezolid-intermediated Enterococcus(LIE) were detected, accounting for 0.4% (28/6924), and 12 Linezolid-resistant Enterococcus(LRE) were detected, with a detection rate of 0.17% (12/6924). Among the LNSE strains, 23 were resistant to genes. The 40 LNSE strains could be divided into 20 different ST types, with ST16 being the main type, accounting for 12.5% (5/40). Conclusion: The detection of LNSE strains was dominated by Enterococcus faecalis, and the main resistance mechanism of LRE strains was carrying the optrA gene, with 23S rRNA gene mutations also contributing to resistance. New resistance gene phenotypes (optrA +/23S rRNA+) emerged. Most LRE cases were sporadic, and clonal dissemination was observed in some strains.

Melanoma Derived Exosomes Amplify Radiotherapy Induced Abscopal Effect via IRF7/I-IFN Axis in Macrophages.

Radiotherapy (RT) can induce tumor regression outside the irradiation field, known as the abscopal effect. However, the detailed underlying mechanisms remain largely unknown. A tumor-bearing mouse model is successfully constructed by inducing both subcutaneous tumors and lung metastases. Single-cell RNA sequencing, immunofluorescence, and flow cytometry are performed to explore the regulation of tumor microenvironment (TME) by RT. A series of in vitro assays, including luciferase reporter, RNA Pulldown, and fluorescent in situ hybridization (FISH) assays, are performed to evaluate the detailed mechanism of the abscopal effect. In addition, in vivo assays are performed to investigate combination therapy strategies for enhancing the abscopal effect. The results showed that RT significantly inhibited localized tumor and lung metastasis progression and improved the TME. Mechanistically, RT promoted the release of tumor-derived exosomes carrying circPIK3R3, which is taken up by macrophages. circPIK3R3 promoted Type I interferon (I-IFN) secretion and M1 polarization via the miR-872-3p/IRF7 axis. Secreted I-IFN activated the JAK/STAT signaling pathway in CD8+ T cells, and promoted IFN-γ and GZMB secretion. Together, the study shows that tumor-derived exosomes promote I-IFN secretion via the circPIK3R3/miR-872-3p/IRF7 axis in macrophages and enhance the anti-tumor immune response of CD8+ T cells.

Delineating the early dissemination mechanisms of acral melanoma by integrating single-cell and spatial transcriptomic analyses.

Acral melanoma (AM) is a rare subtype of melanoma characterized by a high incidence of lymph node (LN) metastasis, a critical factor in tumor dissemination and therapeutic decision-making. Here, we employ single-cell and spatial transcriptomic analyses to investigate the dynamic evolution of early AM dissemination. Our findings reveal substantial inter- and intra-tumor heterogeneity in AM, alongside a highly immunosuppressive tumor microenvironment and complex intercellular communication networks, particularly in patients with LN metastasis. Notably, we identify a strong association between MYC+ Melanoma (MYC+MEL) and FGFBP2+NKT cells with LN metastasis. Furthermore, we demonstrate that LN metastasis requires a metabolic shift towards fatty acid oxidation (FAO) induced by MITF in MYC+MEL cells. Etomoxir, a clinically approved FAO inhibitor, can effectively suppress MITF-mediated LN metastasis. This comprehensive dataset enhances our understanding of LN metastasis in AM, and provides insights into the potential therapeutic targeting for the management of early AM dissemination.

Decoding the metastatic potential and optimal postoperative adjuvant therapy of melanoma based on metastasis score.

Metastasis is a formidable challenge in the prognosis of melanoma. Accurately predicting the metastatic potential of non-metastatic melanoma (NMM) and determining effective postoperative adjuvant treatments for inhibiting metastasis remain uncertain. In this study, we conducted comprehensive analyses of melanoma metastases using bulk and single-cell RNA sequencing data, enabling the construction of a metastasis score (MET score) through diverse machine-learning algorithms. The reliability and robustness of the MET score were validated using various in vitro assays and in vivo models. Our findings revealed a distinct molecular landscape in metastatic melanoma characterized by the enrichment of metastasis-related pathways, intricate cell-cell communication, and heightened infiltration of pro-angiogenic tumor-associated macrophages compared to NMM. Importantly, patients in the high MET score group exhibited poorer prognoses and an immunosuppressive microenvironment, featuring increased infiltration of regulatory T cells and decreased infiltration of CD8+ T cells, compared to the low MET score patient group. Expression of PD-1 was markedly higher in patients with low MET scores. Anti-PD-1 (aPD-1) therapy profoundly affected antitumor immunity activation and metastasis inhibition in these patients. In summary, our study demonstrates the effectiveness of the MET score in predicting melanoma metastatic potential. For patients with low MET scores, aPD-1 therapy may be a potential treatment strategy to inhibit metastasis. Patients with high MET scores may benefit from combination therapies.

Prediction of survival and immunotherapy response by the combined classifier of G protein-coupled receptors and tumor microenvironment in melanoma.

BACKGROUND: Melanoma is the deadliest form of skin tumor, and G protein-coupled receptors (GPCRs) play crucial roles in its carcinogenesis. Furthermore, the tumor microenvironment (TME) affects the overall survival (OS) and the response to immunotherapy. The combination of GPCRs and TME from a multi-omics perspective may help to predict the survival of the melanoma patients and their response to immunotherapy. METHODS: Bulk-seq, single-cell RNA sequencing (scRNA-seq), gene mutations, immunotherapy responses, and clinicopathologic feature data were downloaded from public databases, and prognostic GPCRs and immune cells were screened using multiple machine learning algorithms. The expression levels of GPCRs were detected using real-time quantitative polymerase chain reaction (qPCR) in A375 and HaCaT cell lines. The GPCR-TME classifier was constructed and verified using different cohorts and multi-omics. Gene set enrichment analysis (GSEA), weighted gene co-expression network analysis (WGCNA), and tracking tumor immunophenotype (TIP) were used to identify the key biological pathways among the GPCR-TME subgroups. Then, tumor mutational burden (TMB), vital mutant genes, antigen presentation genes, and immune checkpoints were compared among the subgroups. Finally, the differences in immunotherapy response rates among the GPCR-TME subgroups were investigated. RESULTS: A total of 12 GPCRs and five immune cell types were screened to establish the GPCR-TME classifier. No significant differences in the expression levels of the 12 GPCRs were found in the two cell lines. Patients with high GPCR score or low TME score had a poor OS; thus, the GPCRlow/TMEhigh subgroup had the most favorable OS. The scRNA-seq result revealed that immune cells had a higher GPCR score than tumor and stromal cells. The GPCR-TME classifier acted as an independent prognostic factor for melanoma. GSEA, WGCNA, and TIP demonstrated that the GPCRlow/TMEhigh subgroup was related to the activation and recruitment of anti-tumor immune cells and the positive regulation of the immune response. From a genomic perspective, the GPCRlow/TMEhigh subgroup had higher TMB, and different mutant genes. Ultimately, higher expression levels of antigen presentation genes and immune checkpoints were observed in the GPCRlow/TMEhigh subgroup, and the melanoma immunotherapy cohorts confirmed that the response rate was highest in the GPCRlow/TMEhigh cohort. CONCLUSIONS: We have developed a GPCR-TME classifier that could predict the OS and immunotherapy response of patients with melanoma highly effectively based on multi-omics analysis.

Intratumoral CD73: An immune checkpoint shaping an inhibitory tumor microenvironment and implicating poor prognosis in Chinese melanoma cohorts.

Background: As a novel immune checkpoint, CD73 has been reported to play prominent roles in several malignancies. However, the significance of CD73 in melanoma remains ambiguous. This study sought to reveal the impact of CD73 on the tumor microenvironment (TME) and patients' prognosis, and to investigate whether CD73 could be a therapeutic target in Chinese melanomas, which were dominated by acral and mucosal subtypes. Methods: Two independent Chinese cohorts of 194 patients with melanoma were enrolled. CD73 and PD-L1 expression as well as CD8+ and CD56+ cell infiltrations were evaluated by immunohistochemistry in 194 resected melanoma samples. Clinical outcomes of patients were assessed utilizing the Kaplan-Meier plotter and Cox proportional hazard analysis. RNA-seq data was obtained from TCGA database. Gene set functional annotations were performed based on GO, KEGG and GSEA analysis. CIBERSORT, ssGSEA and TIMER were used to explore the association between CD73 and immune infiltration. These findings were validated by establishing tumor xenograft model, and functions of tumor-infiltrating immune cells were examined by flow cytometry and immunofluorescence. Results: High CD73 expression showed poorer clinical outcomes and was identified as an independent prognostic indicator for survival in two cohorts. Expression of CD73 was more prevalent than PD-L1 in Chinese melanoma cohorts (54.6% vs 23.2%). Co-expression of both immune checkpoints was infrequent (12.9%) in melanoma, and 54.4% of PD-L1 negative cases showed elevated expression of CD73. CD73high tumors showed a microenvironment with fewer CD8+ T cells and CD56+ NK cells infiltration, which displayed a dysfunctional phenotype. With the treatment of CD73 inhibitor APCP, the amount of CD8+ T cells and CD56+ NK cells infiltrated in tumors was elevated and the immunosuppressive effect of CD73 was eliminated. Conclusions: High CD73 expression was associated with an inhibitory TME and adverse clinical outcomes of melanoma. In comparison to PD-L1, CD73 was more prevalent and possessed more definite prognostic significance. Therefore, it may serve as a prognostic indicator and immunotherapeutic target next to PD-L1 in melanoma for Chinese population.

The Effects of Ring Strain on Cyclic Tetraaryl[5]cumulenes.

Cyclic tetraaryl[5]cumulenes (1 a-f) have been synthesized and studied as a function of increasing ring strain. The magnitude of ring strain is approximated by the extent of bending of the cumulenic core as assessed by a combination of X-ray crystallographic analysis and DFT calculations. Trends are observed in 13 C NMR, UV-vis, and Raman spectra associated with ring strain, but the effects are small. In particular, the experimental HOMO-LUMO gap is not appreciably affected by bending of the [5]cumulene framework from ca. 174° (λmax =504 nm) in 1 a to ca. 178° (λmax =494 nm) in 1 f.

Protein tyrosine phosphatase 1B regulates fibroblasts proliferation, motility and extracellular matrix synthesis via the MAPK/ERK signalling pathway in keloid.

Keloid is a fibroproliferative disorder resulting from trauma, characterized by abnormal activation of keloid fibroblasts and excessive deposition of extracellular matrix (ECM). It affects life quality of patients and lacks of effective therapeutic targets. Protein tyrosine phosphatase 1B (PTP1B) belongs to the protein tyrosine phosphatases and participates in many cellular processes such as metabolism, proliferation and motility. It has been reported that PTP1B negatively regulated diabetic wound healing and tumor progression. However, its effects in keloid remain unclear. Here, we aimed to evaluate the effects of PTP1B on keloid fibroblasts which play essential roles in keloids pathogenesis. Our results revealed that PTP1B expression was decreased both in keloid tissues and in keloid fibroblasts compared to healthy controls. Keloid fibroblasts (KFs) showed higher cell proliferation, motility, ECM production and ERK activity than normal fibroblasts (NFs). Overexpression of PTP1B in KFs and NFs inhibited cell proliferation, motility, ECM synthesis and the MAPK/ERK signalling pathway while knockdown of PTP1B showed converse effects. The rescue experiments with ERK inhibitor further verified that MAPK/ERK signalling pathway involved in PTP1B regulatory network. Taken together, our findings indicated that overexpression of PTP1B suppressed keloid fibroblasts bio-behaviours and promoted their phenotype switch to normal cells via inhibiting the MAPK/ERK signalling pathway, suggesting it may be a potential anti-keloid therapy.

PSMC2 knockdown suppressed tumor progression of skin cutaneous melanoma.

Skin cutaneous melanoma (SKCM) is the most lethal tumor among three of the major malignant cancers of the skin. The mechanism underlying the malignant biological behaviors of SKCM is not fully clear. Our study intended to verify the molecular mechanism of proteasome 26 S subunit ATPase 2 (PSMC2) in malignant biological behaviors of SKCM. The Cancer Genome Atlas (TCGA) database was used to analyze the expression of PSMC2 in SKCM and its impact on prognosis. PSMC2 expression in 105 paired SKCM tissues was investigated by immunohistochemistry (IHC), its functional roles were verified using a series of cell experiments, and the underlying pathway was detected by protein-chip technology and gene set enrichment analysis. We found that PSMC2 was significantly upregulated in SKCN patients from TCGA datasets and verified in clinical SKCM tissues. Moreover, high PSMC2 was shown to closely correlate with the pathological stages and lymphatic metastasis of SKCM patients. Functionally, knockdown of PSMC2 suppressed the progression of SKCM through inhibiting cell proliferation, migration, and DNA damage in vitro as well as cell growth in vivo, whereas inducing apoptosis, cycle arrest in G2 phase. Similarly, pharmaceutical inhibition of proteasome with MG132 mimicked the PSMC2 knockdown induced defects in cell cycle arrest, apoptosis and proliferation, while overexpression of PSMC2 has the opposite effects. Mechanistically, the silence of PSMC2 remarkably elevated the pro-apoptotic proteins DR6, IGFBP-4, p21, and p53, while inhibited the anti-apoptosis protein TRAILR-3 and the proteins related to the Wnt signaling pathway. The present study revealed that PSMC2 participated in a positive regulation to promote the progression of SKCM through regulating the Wnt signaling pathway. Our findings may offer a new mechanism underlying the development and progression of SKCM, and a deeper understanding of PSMC2 may contribute to SKCM treatment.

REST promotes ETS1-dependent vascular growth in medulloblastoma.

Expression of the RE1-silencing transcription factor (REST), a master regulator of neurogenesis, is elevated in medulloblastoma (MB) tumors. A cell-intrinsic function for REST in MB tumorigenesis is known. However, a role for REST in the regulation of MB tumor microenvironment has not been investigated. Here, we implicate REST in remodeling of the MB vasculature and describe underlying mechanisms. Using RESTTG mice, we demonstrate that elevated REST expression in cerebellar granule cell progenitors, the cells of origin of sonic hedgehog (SHH) MBs, increased vascular growth. This was recapitulated in MB xenograft models and validated by transcriptomic analyses of human MB samples. REST upregulation was associated with enhanced secretion of proangiogenic factors. Surprisingly, a REST-dependent increase in the expression of the proangiogenic transcription factor E26 oncogene homolog 1, and its target gene encoding the vascular endothelial growth factor receptor-1, was observed in MB cells, which coincided with their localization at the tumor vasculature. These observations were confirmed by RNA-Seq and microarray analyses of MB cells and SHH-MB tumors. Thus, our data suggest that REST elevation promotes vascular growth by autocrine and paracrine mechanisms.

Photo-ammonification in surface water samples: Mechanism and influencing factors.

Dissolved organic nitrogen (DON) accounts for a large proportion of the total aquatic nitrogen. Compared with dissolved inorganic nitrogen (DIN), the reactivity of DON has received limited attention. Photo-ammonification contributes significantly to the transformation of DON to DIN. However, information on the mechanism of this process is limited. This study investigated the photo-ammonification process of different natural surface water samples. The effects of seasons and rainfall on this process were explored, and the contributing factors were identified. Results showed that the seasonal effect on photo-ammonification differed for different water samples, whereas rainfall increased the rates of photo-ammonification for most of the lakes. The concentrations of reactive species, including triplet states of chromophoric dissolved organic matter (3CDOM*) and singlet oxygen (1O2), were found to be significantly correlated with water optical-parameters. Multivariable linear regression analysis (R2 = 0.617) revealed that the photo-ammonification of DON was mainly facilitated by 3CDOM* whereas 1O2 competed with 3CDOM* and showed an inhibiting effect. The components of dissolved organic matter (DOM) were identified by fluorescence excitation emission matrices coupled with parallel factor analysis and were found to be greatly influenced by the location. Allochthonous humic-like components were found to promote the production of reactive species while tryptophan-like component was found to be a reactive species consumer. This study revealed that the composition of DOM and the reactive species governed the rates of photo-ammonification.

Licochalcone A is a Natural Selective Inhibitor of Arginine Methyltransferase 6.

Arginine methylation is a post-translational modification that is implicated in multiple biological functions including transcriptional regulation. The expression of protein arginine methyltransferases (PRMT) has been shown to be upregulated in various cancers. PRMTs have emerged as attractive targets for the development of new cancer therapies. Here, we describe the identification of a natural compound, licochalcone A, as a novel, reversible and selective inhibitor of PRMT6. Since expression of PRMT6 is upregulated in human breast cancers and is associated with oncogenesis, we used the human breast cancer cell line system to study the effect of licochalcone A treatment on PRMT6 activity, cell viability, cell cycle, and apoptosis. We demonstrated that licochalcone A is a non-S-adenosyl L-methionine (SAM) binding site competitive inhibitor of PRMT6. In MCF-7 cells, it inhibited PRMT6-dependent methylation of histone H3 at arginine 2 (H3R2), which resulted in a significant repression of estrogen receptor activity. Licochalcone A exhibited cytotoxicity towards human MCF-7 breast cancer cells, but not MCF-10A human breast epithelial cells, by upregulating p53 expression and blocking cell cycle progression at G2/M, followed by apoptosis. Thus, licochalcone A has potential for further development as a therapeutic agent against breast cancer.

Circular RNA circ_0020710 drives tumor progression and immune evasion by regulating the miR-370-3p/CXCL12 axis in melanoma.

BACKGROUND: Circular RNAs (circRNAs) have been reported to have critical regulatory roles in tumor biology. However, their contribution to melanoma remains largely unknown. METHODS: CircRNAs derived from oncogene CD151 were detected and verified by analyzing a large number of melanoma samples through quantitative real-time polymerase chain reaction (qRT-PCR). Melanoma cells were stably transfected with lentiviruses using circ_0020710 interference or overexpression plasmid, and then CCK-8, colony formation, wound healing, transwell invasion assays, and mouse xenograft models were employed to assess the potential role of circ_0020710. RNA immunoprecipitation, luciferase reporter assay and fluorescence in situ hybridization were used to evaluate the underlying mechanism of circ_0020710. RESULTS: Our findings indicated that circ_0020710 was generally overexpressed in melanoma tissues, and high level of circ_0020710 was positively correlated with malignant phenotype and poor prognosis of melanoma patients. Elevated circ_0020710 promoted melanoma cell proliferation, migration and invasion in vitro as well as tumor growth in vivo. Mechanistically, we found that high level of circ_0020710 could upregulate the CXCL12 expression via sponging miR-370-3p. CXCL12 downregulation could reverse the malignant behavior of melanoma cells conferred by circ_0020710 over expression. Moreover, we also found that elevated circ_0020710 was correlated with cytotoxic lymphocyte exhaustion, and a combination of AMD3100 (the CXCL12/CXCR4 axis inhibitor) and anti-PD-1 significantly attenuated tumor growth. CONCLUSIONS: Elevated circ_0020710 drives tumor progression via the miR-370-3p/CXCL12 axis, and circ_0020710 is a potential target for melanoma treatment.

Phoenixin 20 promotes neuronal mitochondrial biogenesis via CREB-PGC-1α pathway.

Neurodegenerative disorders are dreadful diseases that affect millions of people worldwide. Mitochondrial dysfunction is closely associated with the development of neurodegenerative disorders. Phoenixin 20 is a newly discovered neuropeptide with a pleiotropic effect. This study showed that the presence of Phoenixin 20 promoted neuronal mitochondrial biogenesis in vitro. In cultured neuronal M17 cells, Phoenixin 20 increased the expression of mitochondrial regulators PGC-1α, NRF-1, and TFAM at both mRNA and protein levels. The treatment of Phoenixin 20 increased the ratio of mitochondrial vs nuclear DNA (mtDNA/nDNA) and the multiple mitochondrial gene expression as revealed by increasing mRNA expression of Tomm22, Timm50, Atp5d, Ndufs3, and protein expression of NDUFB8. At a cellular level, Phoenixin 20 promoted mitochondrial respiratory rate and cellular ATP production. Mechanistically, we found that Phoenixin 20 induced the phosphorylation of CREB, which suggests that Phoenixin 20 promoted the activation of the CREB pathway. The blockage of CREB by its selective inhibitor H89 prevented the effect of Phoenixin 20 on mitochondrial regulators and biogenesis. Moreover, the study showed that Phoenixin 20 induced the expression of its tentative receptor GPR173 at the mRNA and protein level, and the silence of GPR173 in neuronal cells ablated all its effect on mitochondrial regulation. Collectively, we showed that Phoenixin 20 promoted neuronal mitochondrial biogenesis via the regulation of CREB-PGC-1α pathway. This study revealed a new role and underlying mechanism of Phoenixin 20 in neuronal cells, suggesting it influences the therapeutic implication of neurodegenerative diseases.

Pharmacologic inhibition of lysine-specific demethylase 1 as a therapeutic and immune-sensitization strategy in pediatric high-grade glioma.

BACKGROUND: Diffuse midline gliomas (DMG), including brainstem diffuse intrinsic pontine glioma (DIPG), are incurable pediatric high-grade gliomas (pHGG). Mutations in the H3 histone tail (H3.1/3.3-K27M) are a feature of DIPG, rendering them therapeutically sensitive to small-molecule inhibition of chromatin modifiers. Pharmacological inhibition of lysine-specific demethylase 1 (LSD1) is clinically relevant but has not been carefully investigated in pHGG or DIPG. METHODS: Patient-derived DIPG cell lines, orthotopic mouse models, and pHGG datasets were used to evaluate effects of LSD1 inhibitors on cytotoxicity and immune gene expression. Immune cell cytotoxicity was assessed in DIPG cells pretreated with LSD1 inhibitors, and informatics platforms were used to determine immune infiltration of pHGG. RESULTS: Selective cytotoxicity and an immunogenic gene signature were established in DIPG cell lines using clinically relevant LSD1 inhibitors. Pediatric HGG patient sequencing data demonstrated survival benefit of this LSD1-dependent gene signature. Pretreatment of DIPG with these inhibitors increased lysis by natural killer (NK) cells. Catalytic LSD1 inhibitors induced tumor regression and augmented NK cell infusion in vivo to reduce tumor burden. CIBERSORT analysis of patient data confirmed NK infiltration is beneficial to patient survival, while CD8 T cells are negatively prognostic. Catalytic LSD1 inhibitors are nonperturbing to NK cells, while scaffolding LSD1 inhibitors are toxic to NK cells and do not induce the gene signature in DIPG cells. CONCLUSIONS: LSD1 inhibition using catalytic inhibitors is selectively cytotoxic and promotes an immune gene signature that increases NK cell killing in vitro and in vivo, representing a therapeutic opportunity for pHGG. KEY POINTS: 1. LSD1 inhibition using several clinically relevant compounds is selectively cytotoxic in DIPG and shows in vivo efficacy as a single agent.2. An LSD1-controlled gene signature predicts survival in pHGG patients and is seen in neural tissue from LSD1 inhibitor-treated mice.3. LSD1 inhibition enhances NK cell cytotoxicity against DIPG in vivo and in vitro with correlative genetic biomarkers.

A novel magnesium-based oxygen releasing compound for eutrophic water remediation.

Eutrophication of surface water bodies is a global problem in recent years. Dosing polluted water with oxygen releasing compounds (ORCs), especially those that can remove excessive nutrients simultaneously is regarded as one of the most economical and eco-friendly methods of treating eutrophic waters. In this study, a novel Mg-based ORC was synthesized and characterized as a magnesium hydroxide and hydrogen peroxide complex (MHHPC) with Mg to H2O2 ratio of 2:1. Oxygen-releasing, pH-adjusting and nutrient-removal potentials of MHHPC were evaluated in nano-pure and eutrophic water. The overall performance of MHHPC in preventing the eutrophic water from turning black and odorous was compared with the performance of other ORCs namely, MgO2, CaO2 and the combination of MgCl2 and H2O2. The results showed that MHHPC was capable of constantly releasing oxygen to aqueous phase over a period of one week. Phosphate and ammonia nitrogen in synthetic buffered water can were removed as struvite and other precipitates from the aqueous phase. In the synthetic eutrophic water, all the ORCs tested were able to reduce aqueous ammonia nitrogen below 0.5 mM, while only CaO2 and MHHPC successfully removed the aqueous phosphate. However, CaO2 and MgCl2+H2O2 significantly inhibited microbial activity.

Telocytes inhibited inflammatory factor expression and enhanced cell migration in LPS-induced skin wound healing models in vitro and in vivo.

BACKGROUND: Cell proliferation and death are key components of wound healing and tissue repair. Telocytes (TCs) represent a newly discovered cell type that can protect tissue from acute injury via cell-cell communication with adjacent cells. The aim of this study was to use a mouse model of skin wound healing and lipopolysaccharide (LPS)-induced cell injury to evaluate the effects of TCs on skin wound healing in vivo and in vitro. MATERIAL/METHODS: Immunohistochemical staining was performed to evaluate the alteration of TCs in tissues from normal and chronic wound patients. Then, a male C57BL/6 mouse wound model of the back was established. The mice were divided randomly into three groups, and wound healing was estimated according to the wound healing rate and histology. An LPS-induced co-culture model of a mouse lung telocyte cell line (TCs) with human keratinocyte (HaCaT), human dermal microvascular endothelial cell (HDMEC) or murine fibroblast (L929) cell lines was established to analyse the effects of TCs on constitutive cell types of the skin. Cell proliferation, migration and apoptosis were examined, and reactive oxygen species (ROS) and inflammatory factors in HaCaT cells, HDMECs, and L929 cells were detected to study the mechanisms involved in TC protection in skin wounds. RESULTS: TCs were significantly increased in tissues from chronic wound patients compared with healthy controls. Wound healing was significantly improved in wound mouse models treated with exogenous TCs compared with LPS-induced models. TCs reversed the LPS-induced inhibition of HaCaT cells and HDMECs and reduced the LPS-induced apoptosis of HaCaT cells and the death ratios of HDMECs and L929 cells. TCs reversed LPS-induced ROS in HDMECs and L929 cells and decreased inflammatory factor mRNA levels in HaCaT cells, HDMECs and L929 cells. CONCLUSIONS: TCs reduce wound healing delay, and inflammatory responses caused by LPS might be mediated by inflammatory inhibition, thus restricting apoptosis and promoting migration of the main component cell types in the skin.

Elevated kindlin-2 promotes tumour progression and angiogenesis through the mTOR/VEGFA pathway in melanoma.

BACKGROUND: In our previous study, kindlin-2 promoted skin wound healing and decreased the permeability of neovascularization during angiogenesis. Herein, we explored the biological function and underlying mechanism of kindlin-2 in cutaneous melanoma. METHODS AND RESULTS: Through a series of in vitro assays, we found that high levels of kindlin-2 promoted migration and invasion of melanoma cells without influencing cell proliferation. Quantitative real-time polymerase chain reaction (qRT-PCR) and western blot analyses showed that upregulated kindlin-2 promoted the cellular epithelial-mesenchymal transition (EMT). Importantly, we found that melanoma cells overexpressing kindlin-2 promoted angiogenesis and VEGFA secretion in vitro and facilitated tumour growth and lung metastasis in vivo. To unveil the underlying mechanism, we conducted Next-generation sequencing (NGS) and differential expression analyses. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that overlapping differentially expressed genes (DEGs) were primarily enriched in the TGF-ß, mTOR and VEGF signalling pathways. Then, we confirmed that the mTOR/VEGFA pathway was activated during the process of kindlin-2-induced melanoma progression and angiogenesis. Moreover, we demonstrated that kindlin-2 was significantly overexpressed in clinical melanoma samples and that a high level of kindlin-2 predicted a poor prognosis. CONCLUSIONS: Taken together, these findings showed that kindlin-2 promotes angiogenesis and tumour progression via the mTOR/VEGFA pathway.

Cysteinyl leukotriene receptor type 1 antagonist montelukast protects against injury of blood-brain barrier.

The blood-brain barrier (BBB) is formed by tightly connected cerebrovascular endothelial cells. Injury of human brain endothelial cells can cause disruption of the BBB and severe injury to brain tissue. Signals mediated cysteinyl leukotrienes (cysLTs) and their receptors are involved in a variety of pathological conditions. In the current study, our results show that oxygen glucose-deprivation/reoxygenation (OGD/R) induced the expression of leukotriene receptor type 1 (cysLT1R) in brain endothelial cells. Blockage of cysLT1R by its specific antagonist montelukast suppressed OGD/R-induced altered permeability of the human brain endothelial cell (EC) monolayer. Mechanistically, montelukast treatment reversed OGD/R-induced reduction of the tight junction proteins occludin and zonula occludens-1 (ZO-1). Montelukast also ameliorated OGD/R-induced reduction of inhibitors of matrix metalloproteinases (TIMPs), such as TIMP-1 and TIMP-2. On the other hand, montelukast suppressed the expression and production of matrix metalloproteinases (MMPs) and cytokines including MMP-2, MMP-9, interleukin 1ß (IL-1ß), tumor necrosis factor-α (TNF-α), and interleukin 6 (IL-6). Using a murine middle cerebral artery occlusion brain injury model, we demonstrated that the administration of montelukast improved the surgery-induced brain injury and protected against disruption of brain endothelial junction proteins such as occludin and ZO-1. Collectively, our data suggest that montelukast might confer protective roles against injury in brain endothelial cells.

Downregulation of RNF128 activates Wnt/ß-catenin signaling to induce cellular EMT and stemness via CD44 and CTTN ubiquitination in melanoma.

BACKGROUND: Ring finger proteins (RNFs) were involved in carcinogenesis. Here, we aimed to explore the detailed mechanism of RNF128 in the progression of melanoma. METHODS: We reanalyzed several gene expression profiles from the Gene Expression Omnibus (GEO) database and obtained the overlapped differential expressed RNF genes. Among them, RNF128 was selected to further explore its expression, the biological significance, and the underlying molecular mechanism, as well as the clinical relevance in melanoma patients. RESULTS: RNF128 was found to be significantly downregulated in the selected datasets, which was further verified in our melanoma tissues. Moreover, RNF128 downregulation was shown to correlate with the malignant phenotype of melanoma, and further functional assays demonstrated that low levels of RNF128 promoted melanoma progression via inducing cell epithelial-mesenchymal transition (EMT) and the acquisition of stemness. Mechanistically, RNF128 interference activated the Wnt pathway via simultaneously ubiquitinating CD44/cortactin (CTTN), resulting in CD44 and c-Myc transcription, thus revealed that RNF128 participated in a positive feedback of the Wnt pathway-CD44 loop. Clinically, we found that patients expressing low RNF128 and high CD44/CTTN levels had a poor prognosis. CONCLUSION: Downregulated RNF128 activates Wnt signaling to induce cellular EMT and stemness by ubiquitinating and degrading CD44/CTTN, and RNF128 is a reliable diagnostic and prognostic biomarker, and a deeper understanding of RNF128 may contribute to the treatment of melanoma.