U937 Cells Research Articles

STUDIES ON GLYCOGEN PHOSPHORYLASE AS A POTENTIAL MOLECULAR TARGET IN GLIOBLASTOMA

Abstract AIMS Glioblastoma (GB) is one of the most aggressive malignancies of the brain and spinal cord. Current standard of care has remained surgical resection of the tumour, followed by radiotherapy and/or chemotherapy. Crucial challenges in the effective treatment of GB include resistance to the main chemotherapeutic, temozolomide. Prognosis is poor with median survival remaining at 14.6 months. A now established hallmark of cancer is the ability of malignant cells to “modify, or reprogram, cellular metabolism”, such as glycolysis upregulation. This hallmark could suggest the potential of targeting a specific metabolic component within glycolysis, thus reduce energy production, resulting in possible apoptosis/suppressed proliferation. Glycogen phosphorylase (GP), an enzyme in glyconeolysis, could be inhibited for a therapeutic effect in GB. In this project, GP levels across the three different isoforms (PYGB, PYGL, and PYGM) and the effect of the GP inhibitor CP-91149 was investigated across three different GB cell lines. METHOD Cell viability of T98G, U251-MG, and U87-MG glioblastoma cell lines and SVGp19 human fetal glial cell line was measured after administration of CP-91149 for 24, 48, and 72 hours. Migration of T98G and U251 cells following treatment with CP-91149 was measured in a wound healing assay. Visualisation of the three GP isoforms was achieved through western blot and confirmed by immunocytochemistry. Flow cytometry was utilized to expand analysis on GP inhibition through assessing cell cycle before and after treatments. RESULTS Results showed CP-91149 had a significant dose-dependent effect in vitro on all cell lines. Inhibition of GP resulted in reduced cell migration, also in a dose-dependent manner. Western blot and immunocytochemistry analysis showed PYGL is the most predominant isoform of GP. CONCLUSION Future work in this project will aim to silence expression of GP in GB cell lines in order to compare the results with that yielded from cells treated with the GP inhibitor CP-91149.

INVESTIGATING CUDC-101 EFFECT AND MECHANISM OF ACTION AGAINST GLIOBLASTOMA IN VITRO

Abstract AIMS Glioblastoma (GB) has complex pathophysiology, difficult treatment and resultant poor prognosis. Its median survival rate is approximately 15 months. The aggressive nature of GB results in rapid disease progression in 60-70% patients often within 12 weeks. Limited treatment options include maximal safe surgical resection, followed by radiotherapy and temozolomide (TMZ). The blood brain barrier restricts chemotherapeutic entry and increases dosage leading to increased side effects and decreased treatment tolerance. Furthermore, significant disease heterogeneity means reoccurrence is expected in most cases, for which there are no standard treatment routes. This coupled with inefficient treatment with temozolomide (TMZ) due to the blood brain barrier, highlights the requirement for novel treatments. METHOD Histone deacetylases (HDAC) and endothelial growth factor receptor (EGFR) are mutated and upregulated in GB allowing tumour infiltration and proliferation. CUDC-101 is known to target both HDAC and EGFR in other cancers making it a promising therapeutic to trial in GB. Cell viability of U251, T98G and U87MG human cells was measured post-CUDC-101 administration at 24, 48 and 72hrs. SVGp19 embryonic non-cancerous human glial cells were used as a control to establish CUDC-101 preliminary specificity. Targeting another major hallmark of GB, wound healing assays were performed to assess CUDC-101 capacity to inhibit migration. In addition to this, long-term cellular resistance was investigated through clonogenic assays. Western blotting was then used to identify non-/treated expression levels of activated EGFR and acetylated histone H3. Expression levels in non/-treated cells were then visualised with fluorescent microscopy. In addition, flow cytometry was utilised to observe cell cycle changes. RESULTS Overall, results indicate CUDC-101 has a dose-dependent effect on cell viability (long-term and short-term) and migration. Additionally, fluorescent images show treatment does increase histone acetylation and decrease total EGFR. CONCLUSION Future work will solidify these datasets and show changes in activated EGFR and acetylated histone H3 using western blotting.

Integrated bioinformatics analysis and experimental validation on malignant progression and immune cell infiltration of LTBP2 in gliomas

BackgroundGliomas are the highly aggressive brain tumor and also the most devastating human tumors. The latent TGF binding proteins (LTBP) had been found to be involved in malignant biological process and could be used as potent biomarkers in several solid tumors. While the role of LTBP family in human glioma remain to be elucidated.MethodsNormalized gene expression and corresponding clinical data of 2407 gliomas samples in public datasets were downloaded from Gliovis. Kaplan–Meier methods and Cox regression analysis was used for survival analyses.Western blot (WB) and Immunohistochemical (IHC) testing were employed to test LTBPs protein level in 154 gliomas samples. Correlation between LTBP2 expression and immune infiltration was evaluated by immunofluorescence (IF) and IHC in glioma tissues. CCK8 and flow cytometric analysis were used to detect the effect of LTBP2 on glioma cells. Orthotopic glioma- mouse models were utilized to evaluate effects in vivo.ResultsLTBP2 mRNA level was dramatically higher in glioma samples compared with non-tumor brain tissues in XENA-TCGA_GTEx, Gill and Gravendeel datasets (all P < 0.01), and its expression positively correlated with glioma WHO grade, IDH1/2 wildtype and mesenchymal subtypes. These results were confirmed by In-house cohort which was detected by WB and IHC. We found that gliomas patients with high LTBP2 level had shorter OS than those with low LTBP2 level. LTBP2 expression significantly associated with glioma immune score (Spearman r = 0.68, P < 0.01)) and strongly correlated with infiltration degreee of macrophages both in lower grade gliomas (LGG) and GBM. Knocking down LTBP2 obviously reduced proliferation and enhanced sensitivity to temozolomide in U87 and U251 cells. Nude mice with lower expression of LTBP2 had slower tumor growth, and accompanied by less tumor-associated macrophages (TAMs) infiltration detected by IHC staining in vivo. Finally, low LTBP2 expression glioma patients who received chemotherapy survived longer than patients with high LTBP2 expression.ConclusionLTBP2 could be used as a prognostic marker, and high LTBP2 expression related to abundant TAMs infiltration and with a worse response to chemotherapy.

Antiproliferative Effects of Naja anchietae and Naja senegalensis Venom Peptides on Glioblastoma Cell Lines.

This study explores the potential of natural bioactive peptides from animal venoms as targeted anti-cancer agents with reduced toxicity. Initially, we screened a broad collection of animal venoms for their antiproliferative activity against cancer cell lines. From this collection, we selected venoms from Naja anchietae and Naja senegalensis due to their promising activity. Utilizing reverse- phase high-performance liquid chromatography (RP HPLC), mass spectrometry (MALDI-TOF MS and MALDI-TOF TOF MSMS), and Edman degradation sequencing, we isolated and characterized three peptides named CTNanc1, CTNanc2, and CTNanc3 from Naja anchietae, and three others named CTNsen1, CTNsen2, and CTNsen3 from Naja senegalensis, each with a molecular weight of around 7 kDa. These purified peptides demonstrated inhibition of U87 glioblastoma cell proliferation, but not of U251 and T98G cells, in cell viability assays. To assess the impact of these treatments on cell viability, apoptosis, and necrosis, flow cytometry assays were conducted on U87 cells at 72 h. The results showed a decrease in cell viability and an increase in dead cells, suggesting that the treatments not only promote apoptosis, but may also lead to increased necrosis or late-stage apoptosis as the exposure time increases. These findings suggest that these peptides could be developed as leads for cancer therapy.

Synthesis of chlorotoxin functionalized metallic nanoparticles and their in vitro evaluation of cytotoxic effects in nervous system cancer cell lines

The treatment of glioblastoma (GB) and neuroblastoma (NB) remains a challenge, as current chemotherapies are plagued with systemic toxicity, drug resistance, and inadequate blood–brain barrier (BBB) penetration. Therefore, novel therapeutic strategies with high specificity and the capacity to bypass the BBB are required. Chlorotoxin (CTX) selectively targets gliomas and neuroectodermal tumors, hence the use of CTX-targeted nanoparticles (NPs) represents a promising therapeutic approach for nervous system (NS) cancers. Bimetallic NPs composed of two metals such as gold-platinum NPs (AuPtNPs) exhibit enhanced anticancer properties compared to single-metal NPs, however their application in studying NS tumors has been relatively limited. CTX-functionalized monometallic gold NPs (CTX-AuNPs) and bimetallic gold-platinum NPs (CTX-AuPtNPs) were synthesized in this study. The NPs were characterized by Ultraviolet-Visible Spectroscopy (UV–vis), Dynamic Light Scattering (DLS), Transmission Electron Microscopy (TEM) and Fourier Transform Infra-Red Spectroscopy (FTIR). Cytotoxicity of NPs was investigated in cancer (U87 and SH-SY5Y) and non-cancer (KMST-6) cells using the water-soluble tetrazolium (WST)-1 assay. The CTX-AuNPs and CTX-AuPtNPs had a core size of ∼5 nm. The CTX-AuPtNPs showed significant anticancer activity in U87 cells possibly due to the synergistic effects of combined metals. Findings obtained from this study demonstrated that CTX can be used to target NS cancers and that bimetallic NPs could be effective in their treatment. More studies are required to investigate the mechanisms of NPs toxicity, and further explore the hyperthermia treatment of NS cancer using the CTX-AuPtNPs.

Mechanism research of Tollip negative feedback regulation in TLR4 signaling pathways based on spinal tuberculosis: Detection of Tollip and NF-κB expression levels

The emergence of drug-resistant mycobacterium tuberculosis (MTB, or TB) strains has led to an increasing incidence of TB. Spinal tuberculosis is the most common extrapulmonary tuberculosis. In the present study, tollip, a negative feedback regulatory factor in TLR4 signaling pathway was chosen based on previous studies on osteoarticular tuberculosis. U937 cells were transfected with recombinant lentivirus containing shRNA (RNA interference, RNAi) or overexpression vector containing Tollip gene and tested in vitro. The expression levels of Tollip and TLR4 were detected by Real-time PCR and immunofluorescence techniques, and the cell morphology and infection effect were observed by DAPI staining. The results suggested that Tollip gene could negatively inhibit the expression of related factors in TLR4 signaling pathway, and thus is a potential biomarker for early diagnosis.

Rs2564978(T) allele associated with severe influenza a disrupts binding site for myeloid differentiation factor PU.1 and reduces &lt;i&gt;CD55/DAF&lt;/i&gt; gene promoter activity in macrophages

An inhibitor of the complement system CD55/DAF is expressed on many cell types. Dysregulation of CD55 expression is associated with increased disease severity during influenza A infection, as well as with vascular complications in pathologies involving excessive activation of the complement system. Using a luciferase reporter system, we performed functional analysis of the single nucleotide polymorphism rs2564978 located in the promoter of the CD55 gene in human pro-monocytic cell line U937. We have shown a decreased activity in activated U937 cells of the CD55 gene promoter carrying minor rs2564978(T) allele associated with the severe course of influenza A(H1N1)pdm09. Using bioinformatic resources, we determined that transcription factor PU.1 can potentially bind to the CD55 promoter region containing rs2564978 in an allele-specific manner. The involvement of PU.1 in modulating CD55 promoter activity was determined by genetic knockdown of PU.1 using small interfering RNAs under specific monocyte activation conditions.

Development and In vitro and In vivo efficacy investigation of multifunctional, targeted, theranostic liposomes for imaging and photodynamic therapy of glioblastoma

The challenge of effectively diagnosing and treating glioblastoma, which has a high incidence and low survival rate, remains unresolved. Consequently, research efforts have increased to develop specific and effective diagnostic and therapeutic agents. In this study, N-acetyl glucosamine-modified, IR780-encapsulated, PEG-coated, nanosized, 68Ga-labeled, neutral, and positively charged theranostic liposomes were developed for GLUT1 targeting for PET/CT and NIR imaging and PDT/PTT of GBM. Characterization, in vitro release profile, stability, radiolabeling efficiency, and labeling stability were evaluated. In vitro cell uptake studies were conducted in RG2 and U87 GBM cell lines, NIR images were obtained in the RG2 and U87 cell lines, and fluorescence microscope images were obtained in the RG2 cell line. PDT/PTT activities were assessed after exposure to NIR light (808 nm, 0.8–1 W/cm2). Cytotoxicity was studied in RG2, U87, and L929 cell lines. In vivo studies were performed in GBM model nude mice. The liposomes had an optimum mean particle size (181–193 nm), high encapsulation efficiency (79–84 %), and zeta potential (−5 to −7 mV). The maximum 68Ga labeling efficiency was achieved with an incubation at 80 °C, pH: 3.5, and 5 min. Liposomes remained stable for 30 days at 4 °C. While the intracellular uptake of targeted formulations was higher than non-targeted ones, it was similar for positively charged and neutral formulations. In U87 and RG2 cell lines, cell viability remained above 50 % for liposomal formulations at a concentration of 1.5 μg/mL exposed to NIR light (808 nm, 0.8–1 W/cm2). The formulations showed significant PDT/PTT activity under NIR light and lower cytotoxicity in the L929 than in RG2 and U87 cell lines. In vivo bioluminescence images and biodistribution studies in U87 tumor-bearing nude mice revealed higher tumor uptake of targeted, IR780-encapsulated liposomal formulations in tumor tissue compared to non-targeted ones. Liposomal formulations showed higher tumor uptake than the free IR780 solution. The data suggest that targeted liposomes have potential as theranostic agents for PDT/PTT and multifunctional glioblastoma imaging.

High loading and sustained-release system of doxorubicin-carbon dots as nanocarriers for cancer therapeutics

Nanocarriers for drugs have been investigated for decades, yet it is still challenging to achieve sustained release from nanomaterials due to drug loading inefficiency and burst release. In this study, we developed novel functional carbon dots (CDs) and investigated the therapeutic efficacy by studying the loading efficiency and release behavior of the anticancer drug doxorubicin (DOX). CDs were successfully synthesized using a one-step pyrolysis method with varying concentrations of citric acid (CA) and thiourea (TU). Functional groups, morphology, particle size, and zeta potential of synthesized CT-CDs and DOX loaded CT-CDs were investigated by UV-visible, Fluorescence, dynamic light scattering, Zeta Potential measurements, FTIR, and transmission electron microscopy. The zeta potential data revealed DOX loading onto CT-CDs by charge difference, i.e. -24.6 ± 0.44 mV (CT-CDs) and 20.57 ± 0.55 mV (DOX-CT-CDs). DOX was loaded on CDs with a loading efficiency of 88.67 ± 0.36%.In vitrodrug release studies confirmed pH-dependent biphasic drug release, with an initial burst effect and sustained release of DOX was found to be 21.42 ± 0.28% (pH 5), 13.30 ± 0.03% (pH 7.4), and 13.95 ± 0.18% (pH 9) even after 144 h at 37 °C. The CT-CDs were non-toxic and biocompatible with L929 Fibroblasts cells. The cytotoxic effect of DOX-CT-CDs showed a concentration-dependent effect after 48 h with Glioblastoma U251 cells. Flow cytometry was used to examine the cellular uptake of CT-CDs and DOX-CT-CDs in L929 and U251 cells. It was observed that the maximum CT-CDs uptake was around 75% at the end of 24 h. This study showed that the synthesized fluorescent CT-CDs demonstrated a high drug loading capacity, pH-dependent sustained release of DOX, and high cellular uptake by mammalian cells. We believe this work provides practical and biocompatible CDs for chemotherapeutic drug delivery that can be applied to other drugs for certain therapeutic aims.

Blocking M2-Like Macrophage Polarization Using Decoy Oligodeoxynucleotide-Based Gene Therapy Prevents Immune Evasion for Pancreatic Cancer Treatment.

M2-like macrophages exhibit immunosuppressive activity and promote pancreatic cancer progression. Reactive oxygen species (ROS) affect macrophage polarization; however, the mechanism remains unclear. This study aimed to elucidate the underlying molecular basis and design a gene therapy to inhibit M2-like polarization. Microarray analysis and immunofluorescence staining were performed in M1-like and M2-like macrophages to ascertain the expression of CYBB, a major intracellular ROS source. Coculture assay and syngeneic orthotopic pancreatic cancer mice models were used to study the mechanism of M2-like skewing. Decoy oligodeoxynucleotides (ODNs) were designed to manipulate CYBB transcription to inhibit M2-like polarization and control tumor growth. Lipopolysaccharide treatment polarized U937 cells to M1-like macrophages in which CYBB expression was increased. In contrast, coculture with PANC-1 cells induced M2-like polarization in U937 cells with CYBB downregulation. High CD204 M2-like expression in combination with low CYBB expression was associated with the worst prognosis in patients with pancreatic cancer. STAT6 and HDAC2 in U937 cells were activated by cancer cell-derived IL4 after coculture and then bound to the CYBB promoter to repress CYBB expression, resulting in M2-like polarization. Diphenyleneiodonium, 8λ³-iodatricyclo[7.4.0.02,⁷]trideca-1(13),2,4,6,9,11-hexaen-8-ylium chloride that inhibits ROS production could block this action. Knockdown of STAT6 and HDAC2 also inhibited M2-like polarization and maintained the M1-like phenotype of U937 cells after coculture. Decoy ODNs interrupting the binding of STAT6 to the CYBB promoter counteracted M2-like polarization and tumor growth and triggered antitumor immunity in vivo. Gene therapy using STAT6-CYBB decoy ODNs can inhibit M2-like polarization, representing a potential therapeutic tool for pancreatic cancer.

Mechanism of Huayu Jiedu Decoction in Inhibiting Malignant Biological Characteristics of Multiple Myeloma

To analyze and explore the effects of Huayu Jiedu Decoction on the malignant biological characteristics of multiple myeloma (MM) cells and its molecular mechanism, so as to provide experimental basis and theoretical basis for the alternative therapy of anti-MM in traditional Chinese medicine. Different concentrations of Huayu Jiedu Decoction were used to intervene myeloma U266 cells. The changes of cell proliferation activity were detected by CCK-8 assay, apoptosis was detected by Annexin V/PI double staining flow cytometry, and apoptosis and protein expression of related signaling pathways were detected by Western blot. Real-time quantitative PCR was used to detect mRNA expression changes of high mobility group protein B1 (HMGB1), CXC chemokine receptor 4 (CXCR4) and interleukin-6 (IL-6). Huayu Jiedu Decoction inhibited the proliferative activity of U266 cells and induced their apoptosis in a concentration and time dependent manner ( r =-0.713, r =-0.827). After treatment with Huayu Jiedu Decoction for 48 h, the expressions of anti-apoptotic protein Bcl-2 and survivin were down-regulated, while the expression of pro-apoptotic protein Bax was up-regulated, and the phosphorylation level of TLR4/NF-κB signaling pathway was inhibited. After intervention of Huayu Jiedu decoction, the expressions of HMGB1 and IL-6 mRNA were significantly decreased, while the expression of CXCR4 was not significantly decreased. Huayu Jiedu Decoction can inhibit the proliferative activity of U266 cells and induce programmed death. Its molecular mechanism may be related to regulating the expression of apoptotic proteins, inhibiting the activation of TLR4/NF-κB pathway and down-regulating the expression of HMGB1 and IL-6 mRNA.

Magnetic lipid-poly(lactic-co-glycolic acid) nanoparticles conjugated with epidermal growth factor receptor antibody for dual-targeted delivery of CPT-11

To entrap sparingly water-soluble drugs like CPT-11 (irinotecan), the poly(lactic-co-glycolic acid) (PLGA) nanoparticle (NP) is highly favored due to its low cytotoxicity and approval for clinical use. On the other hand, entrapping hydrophobic oleic acid-coated iron oxide magnetic nanoparticles (OMNP) in PLGA NP can provide a nanovehicle for magnetically targeted drug delivery. Our goal in this study is to develop a new dual-targeted magnetic lipid-polymer NP for the delivery of CPT-11. We first co-entrap OMNP and CPT-11 in self-assembled lipid-PLGA NP to prepare OLNP@CPT-11. The OLNP@CPT-11 surface was modified with an epidermal growth factor receptor (EGFR) antibody Cetuximab (CET), which can actively target the overexpressed EGFR on the U87 glioblastoma cell surface. The OLNP-CET@CPT-11 enables dual targeting through both external magnetic guidance and CET-mediated active targeting. The NP was characterized for physicochemical properties using various analytical techniques. In vitro study confirms ligand-receptor interaction results in enhanced endocytosis of OLNP-CET@CPT-11 by U87 cells, which offers increased cytotoxicity and elevated cell apoptosis rates. Furthermore, magnetic guidance of OLNP-CET@CPT-11 to U87 cells can induce cell death exclusively in the magnetically targeted zone. The dual-targeted strategy also provides the best therapeutic efficacy against subcutaneously implanted U87 tumors in nude mice with intravenously delivered OLNP-CET@CPT-11.

Cancer Chemotherapeutic Effect of Vernonia Amygdalina Delile on Glioblastoma Brain Cancer Cell.

This study is targeted at assessing the chemotherapy factor of the ethanol extract of Vernonia amygdalina Delile (VAD). U87 glioblastoma cells were treated with extract and a fraction of Vernonia amygdalina Delile (VAD) was harvested from the herbarium. Cytotoxicity was evaluated to determine the IC50 through microscopic observation followed by an MTT assay. Subsequently, flow cytometry with a FACS type was employed to conduct cell cycle and apoptosis analyses. Annexin V/PI and PI markers were used to assess apoptosis and cell cycle progression. The ethanol extract and ethyl acetate fraction of VAD showed promising effects as cancer chemotherapy in glioblastoma cells. The IC50 values for the extract and fraction were notably low, at 37.65 µg/ml and 10.12 µg/ml, respectively, for U87 cells. Analysis of apoptosis using FACS revealed a more pronounced apoptotic effect of the 15 µg/ml fraction of VAD on both early and late apoptosis compared to the 75 µg/ml extract of VAD. Although some differences in cell cycle properties were observed, there were no significant differences in cell cycle analysis between the extract and fraction. These findings underscore the efficacy of VAD's ethanol extract and ethyl acetate fraction as chemotherapeutic agents against U87 cancer cells. The low IC50 values and significant induction of early apoptosis highlight the cytotoxic effects of these treatments on U87 cells.

Effect of JMJD3-IRF4 Signaling Pathway-Mediated Macrophage Polarization on the Malignant Biological Behavior of Multiple Myeloma Cells

To investigate the effect of macrophage polarization mediated by Jumonji domain containing-3 (JMJD3)-interferon regulatory factor 4 (IRF4) signaling pathway on the malignant biological behavior of multiple myeloma (MM) cells. THP-1 monocytes were induced to differentiate into macrophages by phorbol myristate acetate (PMA). THP-1 macrophages were divided into control group (normal culture), M2 induction group ［added recombinant human interleukin (IL) -4, IL-13 proteins］, M2+JMJD3 protein group (added recombinant human IL-4, IL-13 and JMJD3 proteins) and M2+JMJD3 inhibitor group (added recombinant human IL-4, IL-13 proteins and JMJD3 inhibitor), the proportion of CD206+ cells was detected by flow cytometry, the levels of IL-10 and transforming growth factor-β (TGF-β) in the culture supernatant were detected by ELISA assay, the expression levels of arginase-1 (Arg-1), JMJD3 and IRF4 mRNA were detected by real-time quantitative PCR (qRT-PCR), and the expression levels of Arg-1, JMJD3 and IRF4 proteins were detected by Western blot. Correspondingly, human MM cells U266 were cultured with THP-1 macrophage culture supernatant of each group, Methyl thiazolyl tetrazolium (MTT) method and plate colony formation assay were used to detect cell proliferation, cell apoptosis was detected by flow cytometry, Western blot was used to detect the expression levels of apoptosis-promoting protein Bcl-2-associated X protein (Bax) and cleaved caspase-3 in cells, and Transwell assay was used to detect cell migration and invasion. Compared with the control group, the proportion of CD206+ cells in THP-1 macrophages, the mRNA and protein expression levels of Arg-1, JMJD3 and IRF4, and the levels of IL-10 and TGF-β in the cell culture supernatant in M2 induction group were significantly increased (P <0.001), meanwhile, the proliferation activity and the number of clones of U266 cells were significantly increased (P <0.01), the apoptosis rate and the expression levels of apoptosis-promoting protein Bax and cleaved caspase-3 were significantly decreased (P <0.001), the numbers of migrated cells and invasive cells were increased (P <0.001). Compared with M2 induction group, the proportion of CD206+ cells in THP-1 macrophages, the mRNA and protein expression levels of Arg-1, JMJD3 and IRF4, and the levels of IL-10 and TGF-β in the cell culture supernatant in M2+JMJD3 protein group were further increased (P <0.01), meanwhile, the proliferation activity and the number of clones of U266 cells were further increased (P <0.05), the apoptosis rate and the expression levels of apoptosis-promoting protein Bax and cleaved caspase-3 were further decreased (P <0.01), the numbers of migrated cells and invasive cells were further increased (P <0.001); However, the change trends of the above indexes in M2+JMJD3 inhibitor group were opposite to those in M2+JMJD3 protein group. M2 polarization of macrophages mediated by JMJD3-IRF4 signaling pathway can promote the proliferation, migration and invasion of MM cells, and inhibit cell apoptosis.

Berberine Inhibits the Inflammatory Response Induced by Staphylococcus aureus Isolated from Atopic Eczema Patients via the TNF-α/Inflammation/RAGE Pathways.

Atopic eczema patients exhibit high levels of Staphylococcus aureus (S. aureus) skin colonization. S. aureus can stimulate macrophages and the expression of proinflammatory cytokines. Berberine (BBR), an alkaloid, attenuates S. aureus toxin production. This study investigated if BBR suppressed bacterial growth and inflammatory response induced by eczema-patient-derived S. aureus using murine macrophage (RAW 264.7) and human monocyte cell lines (U937). RAW 264.7 and U937 were treated with BBR at different concentrations and stimulated with heat-killed S. aureus (ATCC #33591) or S. aureus derived from severe eczema patients (EC01-EC10), who were undergoing topical steroid withdrawal, for 24 h. TNF-α protein levels were determined by ELISA, gene expression by qRT-PCR, cell cytotoxicity by trypan blue excursion, and reactive oxygen species (ROS) levels by fluorometric assay. BBR showed a bacteriostatic effect in S. aureus (ATCC strain #33591 and clinical isolates (EC01-EC10) and suppressed TNF-α production in RAW 264.7 and U937 cells exposed to heat-killed S. aureus (ATCC and clinical isolates) dose-dependently without any cell cytotoxicity. BBR (20 µg/mL) suppressed >90% of TNF-α production (p < 0.001), downregulated genes involved in inflammatory pathways, and inhibited S. aureus ROS production in U937 and RAW 264.7 cells (p < 0.01). BBR suppresses S. aureus-induced inflammation via inhibition of TNF-α release, ROS production, and expression of key genes involved in the inflammatory pathway.

Two bis-maltol-polyamines: Synthesis, characterization and studies of their palladium(II) complexes exploring their potential anticancer activity

The interest in the antineoplastic and binding properties shown by the bis-maltol polyamine family, particularly Malten and Maltonis, prompted us to study the Pd2+ complexes of these latter from both a biological and metallo-receptor point of view. The Malten-Pd2+ complex can lodge hard species such as Sr2+ in its coordination-driven preorganized pocket, as confirmed by X-ray diffraction. UV–Vis and NMR data showed that Malten-Pd2+ forms even at acidic pH and exists in aqueous solution in a wide range of pH. The mononuclear complex is stable enough not to release Pd2+ in solution for a long period of time (at least one week), thus Malten-Pd2+, similarly to Maltonis-Pd2+, is suitable to be tested in biological analyses. Studies on the U937 cell line revealed that the effect on cell survival reduction induced by Malten is partially lost in Malten-Pd2+, while no differences where monitored between the effects of Maltonis-Pd2+ and Maltonis, suggesting that the availability of free maltol moieties, that is retained in Maltonis-Pd2+, but not in Malten-Pd2+, is crucial to guarantee the biological activity of these compounds.

Inhibition of Mac-1 allows human macrophages to migrate against the direction of shear flow on ICAM-1.

All immune cells must transit from the blood to distal sites such as the lymph nodes, bone marrow, or sites of infection. Blood borne monocytes traffic to the site of inflammation by adhering to the endothelial surface and migrating along endothelial intracellular adhesion molecule 1 (ICAM-1) by their ligand's macrophage 1 antigen (Mac-1) and lymphocyte functional antigen 1 (LFA-1) to transmigrate through the endothelium. Poor patient prognoses in chronic inflammation and tumors have been attributed to the hyper recruitment of certain types of macrophages. Therefore, targeting the binding of ICAM-1 to its respective ligands provides a novel approach to targeting the recruitment of macrophages. To that end, we determined whether the loss of Mac-1 expression could induce this upstream migration behavior by using blocking antibodies against Mac-1 to examine the effects of hydrodynamic flow on the migration of the human macrophage cell line U-937 on ICAM-1 surfaces. Blocking Mac-1 on U-937 cells led to upstream migration against the direction of shear flow on ICAM-1 surfaces. In sum, the ability of macrophages to migrate upstream when Mac-1 is blocked represents a new avenue to precisely control the differentiation, migration, and trafficking of macrophages.

Relaxometric properties and biocompatibility of a novel nanostructured fluorinated gadolinium metal-organic framework.

A novel Gd-MOF based on tetrafluoro-terephthalic acid has been synthesized and its structure has been solved using X-ray single crystal diffraction data. The compound, with the formula [Gd2(F4BDC)3·H2O]·DMF, is isostructural with other Ln-MOFs based on the same ligand and has been recently reported. Its crystals were also reduced to nanometer size by employing acetic acid or cetyltrimethylammonium bromide (CTAB) as a modulator. The relaxometric properties of the nanoparticles were evaluated in solution by measuring 1H T1 and T2 as a function of the applied magnetic field and temperature. The biocompatibility of Gd-MOFs was evaluated on murine microglial BV-2 and human glioblastoma U251 cell lines. In both cell lines, Gd-MOFs do not modify the cell cycle profile or the activation levels of ERK1/2 and Akt, which are protein-serine/threonine kinases that participate in many signal transduction pathways. These pathways are fundamental in the regulation of a large variety of processes such as cell migration, cell cycle progression, differentiation, cell survival, metabolism, transcription, tumour progression and others. These data indicate that Gd-MOF nanoparticles exhibit high biocompatibility, making them potentially valuable for diagnostic and biomedical applications.

Long-term exposure of human U87 glioblastoma cells to polyethylene microplastics: Investigating the potential cancer progression

Precancerous cells are present in all human bodies. Various environmental triggers can promote the development of cancer. Microplastics, an emerging concern, may potentially act as one such trigger, contributing to cancer initiation or progression. Studies have confirmed the presence of microplastics within the human body. This raises concerns about their potential toxicity and health risks. In the present study, we aimed to investigate the impact of polyethylene microplastics (PE-MPs) within the size range of 37–75 microns on glioblastoma cancer cells. Initially, we assessed the short-term effects of six different concentrations of PE-MPs (20 mg/mL, 10 mg/mL, 5 mg/mL, 2.5 mg/mL, 1.25 mg/mL, and 0.62 mg/mL) on the U87 glioblastoma cell line. The results demonstrated that PE-MPs exposure led to an increase in cell proliferation compared to the untreated control group. Based on these findings, we decided to further explore the long-term effects of PE-MPs on U87 cancer cells. To evaluate the long-term effects, U87 glioblastoma cells were continuously exposed to 0.005 g of PE-MPs over an extended period of 26 days. Chronic exposure to PE-MPs significantly increased the proliferative and migratory capacities of U87 cells compared to the unexposed control group. Furthermore, continuous PE-MPs exposure altered the behavior and morphological characteristics of U87 cells. These cells exhibited a propensity to aggregate and form colonies within the culture flask. The formation of spheroid structures was also observed in the PE-MPs-exposed cell population. The results of this research indicate that polyethylene microplastics can promote the progression of glioblastoma cancer.

Comparison of the effects of photodynamic exposure with the use of chlorine E6 on glioblastoma cells of the U251 line and human embryonic kidney cells of the HEK293 line in vitro

Malignant gliomas of the brain are a global medical and social problem with a trend toward a steady increase in morbidity and mortality rates. A method that enables the visual identification of tumor tissue and simultaneously selectively destroys it is photodynamic therapy, which involves the introduction of a photosensitizer (PS) followed by its activation at a certain wavelength of light. The selectivity of the accumulation of PS in the tumor tissue of the malignant gliomas is one of the key issues in the problem of increasing the effectiveness of photodynamic therapy. Objective: to compare the effects of photodynamic exposure using PS chlorin E6 on human glioblastoma (GB) cells of the U251 line and non-malignant human embryonic kidney cells of the HEK293 line. Material and methods. Groups of cell cultures were formed depending on the conditions of cultivation and exogenous influence: 1) control - cultivated in a standard nutrient medium (Modified Eagle's Medium (MEM)) with L-glutamine, 1 mmol of sodium pyruvate, 10% fetal bovine serum) and experimental: 2) cultivated under the conditions of adding chlorin E6 (concentrations 1.0 and 2.0 μg/ml); 3) cultivated on a nutrient medium without the addition of PS and exposed to laser irradiation (LI) (λ=660 nm, power in the range 0.4-0.6 W, dose in the range 10-75 J/cm2, continuous or pulse mode); 4) cultured under conditions of chlorin E6 addition and subsequent exposure to LI (power in the range 0.4-0.6 W, dose in the range 10-75 J/cm2, continuous or pulse mode). After exposure to the specified experimental factors, dynamic observation with microphotographic registration was performed for 24 h, followed by microscopic and micrometric studies (number of viable cells, total number of cells, mitotic index (MI,%)). Results. PS chlorin E6 is incorporated into the cytoplasm of cells of U251 and HEK293 cell lines, the intensity of fluorescence is comparable. Upon exposure to chlorin E6 (1.0 and 2.0 μg/ml), cytodestructive and antimitotic effects are increased in a dose-dependent manner in the culture of human GB cells of the U251 line. The cytodestructive effect of chlorin E6 on cell cultures of the HEK293 line is less pronounced, but the antimitotic effect is comparable in both types of cell cultures. Under the influence of LI, cytodestructive and antimitotic effects increase in a dose-dependent manner in the culture of human GB cells of the U251 line. The level of cytodestructive and antimitotic effects is significantly lower in the cultures of non-neoplastic HEK293 cells. The most significant drop in the mitotic activity of GB U251 cells (~100%) was recorded at the lowest LI dose of 25 J/cm2, power of 0.6 W in pulse mode. For HEK293 cells, the most significant decrease in mitotic activity (~80%) was recorded at LI with a power of 0.6 W and dose of 75 J/cm2 in continuous mode. Under the combined effect of chlorin E6 (1 and 2 μg/ml, pre-incubation of 4 h) and LI in different modes, the viability of tumor cells in U251 culture decreases in a dose-dependent manner; the smallest dose of LI to achieve the maximum cytotoxic effect is 25 J/cm2, with a power of 0.6 W in pulse mode when using chlorin E6 at a concentration of 2 μg/ml. The specified characteristics of photodynamic exposure do not cause irreversible effects in HEK293 cultures (reference cells). Conclusions. An effective mode of photodynamic exposure to achieve a cytodestructive and antimitotic effect in the culture of human GB cells of the U251 line, which is relatively safe for non-malignant cells, has been established: the combined application of a laser irradiation dose of 25 J/cm2, with a power of 0.6 W in pulse mode during the preliminary incubation of the cell culture with chlorin E6 at a concentration of 2 μg/ml for 4 h.