Arginine Citrullination at the C-Terminal Domain Controls RNA Polymerase II Transcription.

The post-translational modification of key residues at the C-terminal domain of RNA polymerase II (RNAP2-CTD) coordinates transcription, splicing, and RNA processing by modulating its capacity to act as a landing platform for a variety of protein complexes. Here, we identify a new modification at the CTD, the deimination of arginine and its conversion to citrulline by peptidyl arginine deiminase 2 (PADI2), an enzyme that has been associated with several diseases, including cancer. We show that, among PADI family members, only PADI2 citrullinates R1810 (Cit1810) at repeat 31 of the CTD. Depletion of PADI2 or loss of R1810 results in accumulation of RNAP2 at transcription start sites, reduced gene expression, and inhibition of cell proliferation. Cit1810 is needed for interaction with the P-TEFb (positive transcription elongation factor b) kinase complex and for its recruitment to chromatin. In this way, CTD-Cit1810 favors RNAP2 pause release and efficient transcription in breast cancer cells.

USP7 deubiquitinates epigenetic reader ZMYND8 to promote breast cancer cell migration and invasion.

The ubiquitin-proteasome system (UPS), which involves E3 ligases and deubiquitinases (DUBs), is critical for protein homeostasis. The epigenetic reader ZMYND8 (zinc finger MYND-type containing 8) has emerged as an oncoprotein, and its protein levels are elevated in various types of cancer, including breast cancer. However, the mechanism by which ZMYND8 protein levels are increased in cancer remains elusive. Although ZMYND8 has been reported to be regulated by the E3 ligase FBXW7, it is still unknown whether ZMYND8 could be modulated by DUBs. Here, we identified USP7 (ubiquitin carboxyl-terminal hydrolase 7) as a bona fide DUB for ZMYND8. Mechanically, USP7 directly binds to the PBP (PHD-BRD-PWWP) domain of ZMYND8 via its TRAF (tumor necrosis factor receptor-associated factor) domain and UBL (ubiquitin-like) domain and removes F-box and WD repeat domain containing 7 (FBXW7)-catalyzed poly-ubiquitin chains on lysine residue 1034 (K1034) within ZMYND8, thereby stabilizing ZMYND8 and stimulating the transcription of ZMYND8 target genes ZEB1 (zinc finger E-box binding homeobox 1) and VEGFA (Vascular Endothelial Growth Factor A). Consequently, USP7 enhances the capacity of breast cancer cells for migration and invasion through antagonizing FBXW7-mediated ZMYND8 degradation. Importantly, the protein levels of USP7 positively correlates with those of ZMYND8 in breast cancer tissues. These findings delineate an important layer of migration and invasion regulation by the USP7-ZMYND8 axis in breast cancer cells.

Purinergic signaling promotes premature senescence.

Extracellular ATP activates P2 purinergic receptors. Whether purinergic signaling is functionally coupled to cellular senescence is largely unknown. We find that oxidative stress induced release of ATP and caused senescence in human lung fibroblasts. Inhibition of P2 receptors limited oxidative stress-induced senescence, while stimulation with exogenous ATP promoted premature senescence. Pharmacological inhibition of P2Y11 receptor (P2Y11R) inhibited premature senescence induced by either oxidative stress or ATP, while stimulation with a P2Y11R agonist was sufficient to induce cellular senescence. Our data show that both extracellular ATP and a P2Y11R agonist induced calcium (Ca++) release from the endoplasmic reticulum (ER) and that either inhibition of phospholipase C or intracellular Ca++ chelation impaired ATP-induced senescence. We also find that Ca++ that was released from the ER, following ATP-mediated activation of phospholipase C, entered mitochondria in a manner dependent on P2Y11R activation. Once in mitochondria, excessive Ca++ promoted the production of reactive oxygen species in a P2Y11R-dependent fashion, which drove development of premature senescence of lung fibroblasts. Finally, we show that conditioned medium derived from senescent lung fibroblasts, which were induced to senesce through the activation of ATP/P2Y11R-mediated signaling, promoted the proliferation of triple-negative breast cancer cells and their tumorigenic potential by secreting amphiregulin. Our study identifies the existence of a novel purinergic signaling pathway that links extracellular ATP to the development of a protumorigenic premature senescent phenotype in lung fibroblasts that is dependent on P2Y11R activation and ER-to-mitochondria calcium signaling.

A progesterone derivative linked to a stable phospholipid activates breast cancer cell response without leaving the cell membrane.

In hormone-responsive breast cancer cells, progesterone (P4) has been shown to act via its nuclear receptor (nPR), a ligand-activated transcription factor. A small fraction of progesterone receptor is palmitoylated and anchored to the cell membrane (mbPR) forming a complex with estrogen receptor alpha (ERα). Upon hormone exposure, either directly or via interaction with ERα, mbPR activates the SRC/RAS/ERK kinase pathway leading to phosphorylation of nPR by ERK. Kinase activation is essential for P4 gene regulation, as the ERK and MSK1 kinases are recruited by the nPR to its genomic binding sites and trigger chromatin remodeling. An interesting open question is whether activation of mbPR can result in gene regulation in the absence of ligand binding to intracellular progesterone receptor (iPR). This matter has been investigated in the past using P4 attached to serum albumin, but the attachment is leaky and albumin can be endocytosed and degraded, liberating P4. Here, we propose a more stringent approach to address this issue by ensuring attachment of P4 to the cell membrane via covalent binding to a stable phospholipid. This strategy identifies the actions of P4 independent from hormone binding to iPR. We found that a membrane-attached progestin can activate mbPR, the ERK signaling pathway leading to iPR phosphorylation, initial gene regulation and entry into the cell cycle, in the absence of detectable intracellular progestin.

The suppression of cell motility through the reduction of FAK activity and expression of cell adhesion proteins by hAMSCs secretome in MDA-MB-231 breast cancer cells.

Breast cancer is a leading cause of death in women worldwide. Cancer therapy based on stem cells is considered as a novel and promising platform. In the present study, we explore the therapeutic effects of human amniotic mesenchymal stromal cells (hAMSCs) through the reduction of focal adhesion kinase (FAK) activity, SHP-2, and cell adhesion proteins such as Paxillin, Vinculin, Fibronectin, Talin, and integrin αvß3 expression in MDA-MB-231 breast cancer cells. For this purpose, we employed a co-culture system using 6-well plate transwell. After 72 h, hAMSCs-treated MDA-MB-231 breast cancer cells, the activity of focal adhesion kinase (FAK) and the expression of SHP-2 and cell adhesion proteins such as Paxillin, Vinculin, Fibronectin, Talin, and integrin αvß3 expression were analyzed using western blot. The shape and migration of cells were also analyzed. Based on our results, a significant reduction in tumor cell motility through downregulation of the tyrosine phosphorylation level of FAK (at Y397 and Y576/577 sites) and cell adhesion expression in MDA-MB-231 breast cancer cells was demonstrated. Our findings indicate that hAMSCS secretome has therapeutic effects on cancer cell migration through downregulation of FAK activity and expression of cell adhesion proteins.

Characterization, modes of interactions with DNA/BSA biomolecules and anti-tumor activity of newly synthesized dinuclear platinum(II) complexes with pyridazine bridging ligand.

Platinum-based drugs are widely recognized efficient anti-tumor agents, but faced with multiple undesirable effects. Here, four dinuclear platinum(II) complexes, [{Pt(1,2-pn)Cl}2(µ-pydz)]Cl2 (C1), [{Pt(ibn)Cl}2(µ-pydz)]Cl2 (C2), [{Pt(1,3-pn)Cl}2(µ-pydz)]Cl2 (C3) and [{Pt(1,3-pnd)Cl}2(µ-pydz)]Cl2 (C4), were designed (pydz is pyridazine, 1,2-pn is ( ±)-1,2-propylenediamine, ibn is 1,2-diamino-2-methylpropane, 1,3-pn is 1,3-propylenediamine, and 1,3-pnd is 1,3-pentanediamine). Interactions and binding ability of C1-C4 complexes with calf thymus DNA (CT-DNA) has been monitored by viscosity measurements, UV-Vis, fluorescence emission spectroscopy and molecular docking. Binding affinities of C1-C4 complexes to the bovine serum albumin (BSA) has been monitored by fluorescence emission spectroscopy. The tested complexes exhibit variable cytotoxicity toward different mouse and human tumor cell lines. C2 shows the most potent cytotoxicity, especially against mouse (4T1) and human (MDA-MD468) breast cancer cells in the dose- and time-dependent manner. C2 induces 4T1 and MDA-MD468 cells apoptosis, further documented by the accumulation of cells at sub-G1 phase of cell cycle and increase of executive caspase 3 and caspase 9 levels in 4T1 cells. C2 exhibits anti-proliferative effect through the reduction of cyclin D3 and cyclin E expression and elevation of inhibitor p27 level. Also, C2 downregulates c-Myc and phosphorylated AKT, oncogenes involved in the control of tumor cell proliferation and death. In order to measure the amount of platinum(II) complexes taken up by the cells, the cellular platinum content were quantified. However, C2 failed to inhibit mouse breast cancer growth in vivo. Chemical modifications of tested platinum(II) complexes might be a valuable approach for the improvement of their anti-tumor activity, especially effects in vivo.

Effect of substituents at the C3´, C3´N, C10 and C2-meta-benzoate positions of taxane derivatives on their activity against resistant cancer cells.

We tested the effect of substituents at the (1) C3´, C3´N, (2) C10, and (3) C2-meta-benzoate positions of taxane derivatives on their activity against sensitive versus counterpart paclitaxel-resistant breast (MCF-7) and ovarian (SK-OV-3) cancer cells. We found that (1) non-aromatic groups at both C3´ and C3´N positions, when compared with phenyl groups at the same positions of a taxane derivative, significantly reduced the resistance of ABCB1 expressing MCF-7/PacR and SK-OV-3/PacR cancer cells. This is, at least in the case of the SB-T-1216 series, accompanied by an ineffective decrease of intracellular levels in MCF-7/PacR cells. The low binding affinity of SB-T-1216 in the ABCB1 binding cavity can elucidate these effects. (2) Cyclopropanecarbonyl group at the C10 position, when compared with the H atom, seems to increase the potency and capability of the derivative in overcoming paclitaxel resistance in both models. (3) Derivatives with fluorine and methyl substituents at the C2-meta-benzoate position were variously potent against sensitive and resistant cancer cells. All C2 derivatives were less capable of overcoming acquired resistance to paclitaxel in vitro than non-substituted analogs. Notably, fluorine derivatives SB-T-121205 and 121,206 were more potent against sensitive and resistant SK-OV-3 cells, and derivatives SB-T-121405 and 121,406 were more potent against sensitive and resistant MCF-7 cells. (4) The various structure-activity relationships of SB-T derivatives observed in two cell line models known to express ABCB1 favor their complex interaction not based solely on ABCB1.

Comparative characterisation of an ecto-5'-nucleotidase (CD73) in non-tumoral MCF10-A breast cells and triple-negative MDA-MB-231 breast cancer cells.

Ecto-5'-nucleotidase (CD73) hydrolyses 5'AMP to adenosine and inorganic phosphate. Breast cancer cells (MDA-MB-231) express high CD73 levels, and this enzyme has been found to play a tumour-promoting role in breast cancer. However, no studies have sought to investigate whether CD73 has differential affinity or substrate preferences between noncancerous and cancerous breast cells. In the present study, we aimed to biochemically characterise ecto-5'-nucleotidase in breast cancer cell lines and assess whether its catalytic function and tumour progression are correlated in breast cancer cells. The results showed that compared to nontumoral breast MCF-10A cells, triple-negative breast cancer MDA-MB-231 cells had a higher ecto-5'-nucleotidase expression level and enzymatic activity. Although ecto-5'-nucleotidase activity in the MDA-MB-231 cell line showed no selectivity among monophosphorylated substrates, 5'AMP was preferred by the MCF-10A cell line. Compared to the MCF-10A cell line, the MDA-MB-231 cell line has better hydrolytic ability, lower substrate affinity, and high inhibitory potential after treatment with a specific CD73 inhibitor α,ß­methylene ADP (APCP). Therefore, we demonstrated that a specific inhibitor of the ecto-5-nucleotidase significantly reduced the migratory and invasive capacity of MDA-MB-231 cells, suggesting that ecto-5-nucleotidase activity might play an important role in metastatic progression.

Phytofabrication of biocompatible zinc oxide nanoparticle using Gymnema sylvestre and its potent in vitro antibacterial, antibiofilm, and cytotoxicity against human breast cancer cells (MDA-MB-231).

The increasing incidence of breast cancer and bacterial biofilm in medical devices significantly heightens global mortality and morbidity, challenging synthetic drugs. Consequently, greener-synthesized nanomaterials have emerged as a versatile alternative for various biomedical applications, offering new therapeutic avenues. This study explores the synthesis of biocompatible zinc oxide (ZnONPs) nanoparticles using Gymnema sylvestre and its antibacterial, antibiofilm, and cytotoxic properties. Characterization of ZnONPs inferred that UV-Vis spectra exhibited a sharp peak at 370 nm. Fourier transform infrared spectroscopical analysis revealed the presence of active functional groups such as aldehyde, alkyne, cyclic alkene, sulfate, alkyl aryl ether, and Zn-O bonds. X-ray diffraction analysis results confirmed the crystalline nature of the nanoparticle. Scanning electron microscope analysis evidenced hexagonal morphology, and energy-dispersive X-ray analysis confirmed zinc content. High-resolution transmission electron microscope analysis showed hexagonal and rod-shaped ZnONPs with a size of 5 nm. Zeta potential results affirmed the stability of nanoparticles. The ZnONPs effectively inhibited gram-positive (18-20 mm) than gram-negative (12-18 mm) bacterial pathogens with lower bacteriostatic and higher bactericidal values. Biofilm inhibitory property inferred ZnONPs were more effective against gram-positive (38-94%) than gram-negative bacteria (27-86%). The concentration of ZnONPs to exert 50% biofilm-inhibitory is lower against gram-positive bacteria (179.26-203.95 µg/mL) than gram-negative bacteria (201.46-236.19 µg/mL). Microscopic visualization inferred that at 250 µg/mL, ZnONPs strongly disrupted biofilm formation, as evidenced by decreased biofilm density and altered architecture. The cytotoxicity of ZnONPs against breast cancer cells showed a dose-dependent reduction in cell viability with an IC50 value of 19.4 µg/mL. AO/EB staining indicated early and late apoptotic cell death of breast cancer cells under fluorescence microscopy. The results of hemolytic activity validated the biocompatibility of the ZnONPs. Thus, the unique properties of the green-synthesized ZnONPs suggest their potential as effective drug carriers for targeted delivery in cancer therapy and the treatment of biofilm-related infections.

Leptin Promotes Vasculogenic Mimicry in Breast Cancer Cells by Regulating Aquaporin-1.

Leptin is an obesity-related hormone that plays an important role in breast cancer progression. Vasculogenic mimicry (VM) refers to the formation of vascular channels lined by tumor cells. This study aimed to investigate the relationship between leptin and VM in human breast cancer cells. VM was measured by a 3D culture assay. Signal transducers and activators of transcription 3 (STAT3) signaling, aquaporin-1 (AQP1), and the expression of VM-related proteins, including vascular endothelial cadherin (VE-cadherin), twist, matrix metalloproteinase-2 (MMP-2), and laminin subunit 5 gamma-2 (LAMC2), were examined by Western blot. AQP1 mRNA was analyzed by a reverse transcriptase-polymerase chain reaction (RT-PCR). Leptin increased VM and upregulated phospho-STAT3, VE-cadherin, twist, MMP-2, and LAMC2. These effects were inhibited by the leptin receptor-blocking peptide, Ob-R BP, and the STAT3 inhibitor, AG490. A positive correlation between leptin and AQP1 mRNA was observed and was confirmed by RT-PCR. Leptin upregulated AQP1 expression, which was blocked by Ob-R BP and AG490. AQP1 overexpression increased VM and the expression of VM-related proteins. AQP1 silencing inhibited leptin-induced VM and the expression of VM-related proteins. Thus, these results showed that leptin facilitates VM in breast cancer cells via the Ob-R/STAT3 pathway and that AQP1 is a key mediator in leptin-induced VM.

Antioxidant Properties of Lippia alba Essential Oil: A Potential Treatment for Oxidative Stress-Related Conditions in Plants and Cancer Cells.

Lippia alba (Mill.) N.E.Br. ex Britton and P. Wilson is used in folk medicine of Central and South America for its biological activities: i.e., antifungal, antibacterial, antiviral, and anti-inflammatory. Based on ethnopharmacological information and the increasing interest in this species, this work aimed to test a possible wide use of its essential oil (EO) in pharmaceutical and horticultural applications. Therefore, we focused the attention on the antioxidant activity of the oil as a possible tool to overcome the oxidative stress in both applications. For this purpose, we have chosen three aggressive breast cancer cell lines and two horticultural species (Solanum lycopersicum L. and Phaseolus acutifolius L.) that are very sensitive to salt stress. We determined the antioxidant activity of L. alba EO through the quantification of phenols and flavonoids. Regarding tomato and bean plants under salt stress, L. alba EO was used for the first time as a seed priming agent to enhance plant salt tolerance. In this case, the seed treatment enhanced the content of phenolic compounds, reduced power and scavenger activity, and decreased membrane lipid peroxidation, thus mitigating the oxidative stress induced by salt. While in breast cancer cells the EO treatment showed different responses according to the cell lines, i.e., in SUM149 and MDA-MB-231 the EO decreased proliferation and increased antioxidant activity and lipid peroxidation, showing high cytotoxic effects associated with the release of lactate dehydrogenase, vice versa no effect was observed in MDA-MB-468. Such antioxidant activity opens a new perspective about this essential oil as a possible tool to counteract proliferation in some cancer cell lines and in horticulture as a seed priming agent to protect from oxidative damage in crops sensitive to salinity.

Utility of 1.5 Tesla MRI Scanner in the Management of Small Sample Sizes Driven from 3D Breast Cell Culture.

The aim of this work was to use and optimize a 1.5 Tesla magnetic resonance imaging (MRI) system for three-dimensional (3D) images of small samples obtained from breast cell cultures in vitro. The basis of this study was to design MRI equipment to enable imaging of MCF-7 breast cancer cell cultures (about 1 million cells) in 1.5 and 2 mL glass tubes and/or bioreactors with an external diameter of less than 20 mm. Additionally, the development of software to calculate longitudinal and transverse relaxation times is described. Imaging tests were performed using a clinical MRI scanner OPTIMA 360 manufactured by GEMS. Due to the size of the tested objects, it was necessary to design additional receiving circuits allowing for the study of MCF-7 cell cultures placed in glass bioreactors. The examined sample's volume did not exceed 2.0 mL nor did the number of cells exceed 1 million. This work also included a modification of the sequence to allow for the analysis of T1 and T2 relaxation times. The analysis was performed using the MATLAB package (produced by MathWorks). The created application is based on medical MR images saved in the DICOM3.0 standard which ensures that the data analyzed are reliable and unchangeable in an unintentional manner that could affect the measurement results. The possibility of using 1.5 T MRI systems for cell culture research providing quantitative information from in vitro studies was realized. The scanning resolution for FOV = 5 cm and the matrix was achieved at a level of resolution of less than 0.1 mm/pixel. Receiving elements were built allowing for the acquisition of data for MRI image reconstruction confirmed by images of a phantom with a known structure and geometry. Magnetic resonance sequences were modified for the saturation recovery (SR) method, the purpose of which was to determine relaxation times. An application in MATLAB was developed that allows for the analysis of T1 and T2 relaxation times. The relaxation times of cell cultures were determined over a 6-week period. In the first week, the T1 time value was 1100 ± 40 ms, which decreased to 673 ± 59 ms by the sixth week. For T2, the results were 171 ± 10 ms and 128 ± 12 ms, respectively.

Changes in the Sensitivity of MCF-7 and MCF-7/DX Breast Cancer Cells to Cytostatic in the Presence of Metformin.

Multidrug resistance is a serious problem in modern medicine and the reason for the failure of various therapies. A particularly important problem is the occurrence of multidrug resistance in cancer therapies which affects many cancer patients. Observations on the effect of metformin-a well-known hypoglycemic drug used in the treatment of type 2 diabetes-on cancer cells indicate the possibility of an interaction of this substance with drugs already used and, as a result, an increase in the sensitivity of cancer cells to cytostatics. The aim of this study was to evaluate the effect of metformin on the occurrence of multidrug resistance of breast cancer cells. The MCF-7-sensitive cell line and the MCF-7/DX cytostatic-resistant cell line were used for this study. WST-1 and LDH assays were used to evaluate the effects of metformin and doxorubicin on cell proliferation and viability. The effect of metformin on increasing the sensitivity of MCF-7 and MCF-7/DX cells to doxorubicin was evaluated in an MDR test. The participation of metformin in increasing the sensitivity of resistant cells to the effect of the cytostatic (doxorubicin) has been demonstrated.

Exploring the Antitumor Efficacy of N-Heterocyclic Nitrilotriacetate Oxidovanadium(IV) Salts on Prostate and Breast Cancer Cells.

The crystal structures of two newly synthesized nitrilotriacetate oxidovanadium(IV) salts, namely [QH][VO(nta)(H2O)](H2O)2 (I) and [(acr)H][VO(nta)(H2O)](H2O)2 (II), were determined. Additionally, the cytotoxic effects of four N-heterocyclic nitrilotriacetate oxidovanadium(IV) salts-1,10-phenanthrolinium, [(phen)H][VO(nta)(H2O)](H2O)0.5 (III), 2,2'-bipyridinium [(bpy)H][VO(nta)(H2O)](H2O) (IV), and two newly synthesized compounds (I) and (II)-were evaluated against prostate cancer (PC3) and breast cancer (MCF-7) cells. All the compounds exhibited strong cytotoxic effects on cancer cells and normal cells (HaCaT human keratinocytes). The structure-activity relationship analysis revealed that the number and arrangement of conjugated aromatic rings in the counterion had an impact on the antitumor effect. The compound (III), the 1,10-phenanthrolinium analogue, exhibited the greatest activity, whereas the acridinium salt (II), with a different arrangement of three conjugated aromatic rings, showed the lowest toxicity. The increased concentrations of the compounds resulted in alterations to the cell cycle distribution with different effects in MCF-7 and PC3 cells. In MCF-7 cells, compounds I and II were observed to block the G2/M phase, while compounds III and IV were found to arrest the cell cycle in the G0/G1 phase. In PC3 cells, all compounds increased the rates of cells in the G0/G1 phase.

Carbon Nanotube-Mediated Delivery of PTEN Variants: In Vitro Antitumor Activity in Breast Cancer Cells.

Phosphatase and tensin homologue deleted on chromosome 10 (PTEN) is a crucial tumor suppressor protein with frequent mutations and alterations. Although protein therapeutics are already integral to numerous medical fields, their potential remains nascent. This study aimed to investigate the impact of stable, unphosphorylated recombinant human full-length PTEN and its truncated variants, regarding their tumor suppression activity with multiwalled-carbon nanotubes (MW-CNTs) as vehicles for their delivery in breast cancer cells (T-47D, ZR-75-1, and MCF-7). The cloning, overexpression, and purification of PTEN variants were achieved from E. coli, followed by successful binding to CNTs. Cell incubation with protein-functionalized CNTs revealed that the full-length PTEN-CNTs significantly inhibited cancer cell growth and stimulated apoptosis in ZR-75-1 and MCF-7 cells, while truncated PTEN fragments on CNTs had a lesser effect. The N-terminal fragment, despite possessing the active site, did not have the same effect as the full length PTEN, emphasizing the necessity of interaction with the C2 domain in the C-terminal tail. Our findings highlight the efficacy of full-length PTEN in inhibiting cancer growth and inducing apoptosis through the alteration of the expression levels of key apoptotic markers. In addition, the utilization of carbon nanotubes as a potent PTEN protein delivery system provides valuable insights for future applications in in vivo models and clinical studies.

The inhibitory effect of Gremlin-2 on adipogenesis suppresses breast cancer cell growth and metastasis.

BACKGROUND: Gremlin-1 (GREM1) and Gremlin-2 (GREM2) are bone morphogenetic protein antagonists that play important roles in organogenesis, tissue differentiation, and tissue homeostasis. Although GREM1 has been reported to be involved in promoting various cancers, little has been reported about effects of GREM2 on cancer. Recently, it has been reported that GREM2 can inhibit adipogenesis in adipose-derived stromal/stem cells. However, as an inhibitor of adipogenesis, the role of GREM2 in cancer progression is not well understood yet. METHODS: Pre-adipocyte 3T3-L1 cells overexpressing mock or Grem2 were established using a lentiviral transduction system and differentiated into adipocytes-mock and adipocytes-Grem2, respectively. To investigate the effect of adipocyte-Grem2 on breast cancer cells, we analyzed the proliferative and invasion abilities of spheroids using a 3D co-culture system of breast cancer cells and adipocytes or conditioned medium (CM) of adipocytes. An orthotopic breast cancer mouse model was used to examine the role of adipocytes-Grem2 in breast cancer progression. RESULTS: Grem2 overexpression suppressed adipogenesis of 3T3-L1 cells. Proliferative and invasion abilities of spheroids formed by co-culturing MTV/TM-011 breast cancer cells and adipocytes-Grem2 were significantly reduced compared to those of spheroids formed by co-culturing MTV/TM-011 cells and adipocytes-mock. Compared to adipocytes-mock, adipocytes-Grem2 showed decreased mRNA expression of several adipokines, notably IL-6. The concentration of IL-6 in the CM of these cells was also decreased. Proliferative and invasive abilities of breast cancer cells reduced by adipocytes-Grem2 were restored by IL-6 treatment. Expression levels of vimentin, slug, and twist1 in breast cancer cells were decreased by treatment with CM of adipocytes-Grem2 but increased by IL-6 treatment. In orthotopic breast cancer mouse model, mice injected with both MTV/TM-011 cells and adipocytes-Grem2 showed smaller primary tumors and lower lung metastasis than controls. However, IL-6 administration increased both the size of primary tumor and the number of metastatic lung lesions, which were reduced by adipocytes-Grem2. CONCLUSIONS: Our study suggests that GREM2 overexpression in adipocytes can inhibit adipogenesis, reduce the expression and secretion of several adipokines, including IL-6, and ultimately inhibit breast cancer progression.

Metformin ameliorates BMP2 induced adipocyte-like property in breast cancer cells.

Neighboring adipocytes of tumor cells/cancer associated adipocytes supply many factors and fatty acids as fuel to cancer cells for inducing cancer progression and development. Epithelial breast cancer cells also differentiate into several cell types to meet various demands. This study reports that breast cancer cells exhibit inherent adipocyte-like property which is further enhanced in presence of BMP2. Antidiabetic metformin inhibits BMP2 induced adipocyte-like potential in breast cancer cells. Interestingly, breast cancer cells not only show lipid accumulation but also have ability to release lipid content. Thus, this study centers around the presence of the adipocyte cell-like property in breast cancer cells, the significance of BMP2 and metformin that may be explored in designing therapeutics against breast cancer.

Establishing a new model of cancer neuropathic pain in rats.

To guide the treatment of malignant neuropathic pain (MNP) in clinical practice, by inoculating MADB-106 breast cancer cells into the right L4 nerve root in Sprague-Dawley rats, a rat model of MNP was established, providing basic conditions for the study of neuropathic pain and development and application of therapeutic drugs. As the tumor grew over time, it pressed the nerve roots, causing nerve damage. The spinal nerve ligation (SNL) model, which is a neuropathic pain model widely used in rats, was compared with the L4 nerve root SNL model, and histologic examination of the nerve tissue of both models was performed by electron microscopy. In addition to the infiltration and erosion of the L4 nerve by tumor cells, the tumor tissue gradually grew and compressed the L4 nerve roots, resulting in hyperalgesia of the rat's posterior foot on the operative side. Some spontaneous pain phenomena were also observed, such as constant lifting or licking of the posterior foot on the operative side under quiet conditions. Electron microscopy images showed that nerve injury was due to progressive compression by the tumor, cells of which were visualized, but the injury was lighter than that in SNL rats. Imaging showed a paravertebral tumor near the L4 nerve root in the carcinomatous neuropathic pain model rat. These results suggest that progressive compression of the nerve by a malignant tumor leads to nerve damage similar to the behavioral changes associated with chronic compression injury resulting from a loose ligature of the nerve. The cancer neuropathologic pain model at the L4 nerve root was successfully established in Sprague-Dawley rats.

Anti-Migratory and Cytotoxic Activities of [Ga(8-hydroxyquinolinato)3 ]: Roles of Endogenous Cu(II) and Drug-Induced Phenotypic Changes.

As shown by IncuCyte Zoom imaging proliferation assays, invasive triple-negative human breast MDA-MB-231 cancer cells treated with sub-toxic doses (5.0-20 µM, 72 h) of [GaQ3 ] (Q=8-hydroxyquinolinato) caused profound morphological changes and inhibition of cell migration, which were likely due to terminal cell differentiation or similar phenotypical change. This is the first demonstration of potential use of a metal complex in differentiation anti-cancer therapy. Additionally, a trace amount of Cu(II) (0.20 µM) added to the medium dramatically increased [GaQ3 ] cytotoxicity (IC50 ~2 µM, 72 h) due to its partial dissociation and the action of the HQ ligand as a Cu(II) ionophore, as shown with electrospray mass spectrometry and fluorescence spectroscopy assays in the medium. Hence, cytotoxicity of [GaQ3 ] is strongly linked to ligand binding of essential metal ions in the medium, for example, Cu(II). Appropriate delivery mechanisms of such complexes and their ligands could enable a powerful new triple therapeutic approach for cancer chemotherapy, including cytotoxicity against primary tumour, arrest of metastases, and activation of innate and adaptive immune responses.

Downregulation of Pinkbar/pAKT and MMP2/MMP9 Expression in MDA-MB-231 Breast Cancer Cells as Potential Targets in Cancer Therapy by hAMSCs Secretome.

Breast cancer is one of the leading causes of cancer-related deaths among women worldwide. Cancer therapy based on stem cells is considered as a novel and promising platform. In the present study, we explored the therapeutic effects of human amniotic mesenchymal stromal cells (hAMSCs) through Pinkbar (planar intestinal- and kidney-specific BAR domain protein), pAKT, and matrix metalloproteinases including MMP2 and MMP9 on MDA-MB-231 breast cancer cells. For this purpose, we employed a co-culture system using Transwell 6-well plates with a pore size of 0.4 µm. After 72 h, the hAMSCs-treated MDA-MB-231 breast cancer cells, the expression of epidermal growth factor receptor (EGFR), and c-Src (a key mediator in EGFR signaling pathway), Pinkbar, pAKT, MMP2, and MMP9 were analyzed using quantitative real time PCR and western blot methods. Based on 2D and 3D cell culture models, significant reduction of tumor cell growth and motility through downregulation of EGFR, c-Src, Pinkbar, pAKT, MMP2, and MMP9 were found in MDA-MB-231 breast cancer cells. Moreover, induction of cellular apoptosis was also reported. Our finding indicates that the hAMSCS secretome has therapeutic effects on cancer cells. To identify the details of the molecular mechanisms, more experiments will be required.