Toll-like receptor 9 expression is associated with breast cancer sensitivity to the growth inhibitory effects of bisphosphonates in vitro and in vivo.
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SUBMITTER: Sandholm J
PROVIDER: S-EPMC5349995 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:BackgroundNumerous studies have implicated the aryl hydrocarbon receptor (AhR) as a potential therapeutic target for several human diseases, including estrogen receptor alpha (ER?) positive breast cancer. Aminoflavone (AF), an activator of AhR signaling, is currently undergoing clinical evaluation for the treatment of solid tumors. Of particular interest is the potential treatment of triple negative breast cancers (TNBC), which are typically more aggressive and characterized by poorer outcomes. Here, we examined AF's effects on two TNBC cell lines and the role of AhR signaling in AF sensitivity in these model cell lines.MethodsAF sensitivity in MDA-MB-468 and Cal51 was examined using cell counting assays to determine growth inhibition (GI50) values. Luciferase assays and qPCR of AhR target genes cytochrome P450 (CYP) 1A1 and 1B1 were used to confirm AF-mediated AhR signaling. The requirement of endogenous levels of AhR and AhR signaling for AF sensitivity was examined in MDA-MB-468 and Cal51 cells stably harboring inducible shRNA for AhR. The mechanism of AF-mediated growth inhibition was explored using flow cytometry for markers of DNA damage and apoptosis, cell cycle analysis, and ?-galactosidase staining for senescence. Luciferase data was analyzed using Student's T test. Three-parameter nonlinear regression was performed for cell counting assays.ResultsHere, we report that ER?-negative TNBC cell lines MDA-MB-468 and Cal51 are sensitive to AF. Further, we presented evidence suggesting that neither endogenous AhR expression levels nor downstream induction of AhR target genes CYP1A1 and CYP1B1 is required for AF-mediated growth inhibition in these cells. Between these two ER? negative cell lines, we showed that the mechanism of AF action differs slightly. Low dose AF mediated DNA damage, S-phase arrest and apoptosis in MDA-MB-468 cells, while it resulted in DNA damage, S-phase arrest and cellular senescence in Cal51 cells.ConclusionsOverall, this work provides evidence against the simplified view of AF sensitivity, and suggests that AF could mediate growth inhibitory effects in ER?-positive and negative breast cancer cells, as well as cells with impaired AhR expression and signaling. While AF could have therapeutic effects on broader subtypes of breast cancer, the mechanism of cytotoxicity is complex, and likely, cell line- and tumor-specific.
| S-EPMC4037283 | biostudies-literature
Project description:ObjectivesSaliva can suppress osteoclastogenesis, but the underlying mechanism has not been discovered yet. Considering that endotoxins suppress osteoclastogenesis in bone marrow cultures and that saliva contains endotoxins, it was reasonable to hypothesize that the impact of saliva on osteoclastogenesis requires toll-like receptor 4 signaling.Material and methodsTo test this hypothesis, we blocked toll-like receptor 4 signaling with TAK-242 in the presence of saliva in murine bone marrow cultures. Osteoclastogenesis was evaluated based on gene expression analysis and histochemical staining for tartrate-resistant acid phosphatase. Resorption was performed on dentine.ResultsWe report that TAK-242 reversed the inhibitory effect of fresh sterile saliva on the formation of multinucleated cells that stained positive for tartrate-resistant acid phosphatase. In line with this finding, TAK-242 increased the expression of the osteoclast functional genes cathepsin K, calcitonin receptor, and tartrate-resistant acid phosphatase in the presence of saliva. TAK-242 also supported the expression of NFATc1, the master regulator of osteoclastogenesis, as well as DC-STAMP and Atp6v0d2, both being cell fusion genes. In support of the hypothesis, depletion of saliva from endotoxin partially reversed the inhibitory effect on osteoclastogenesis. Moreover, salivary pellicle on plastic and titanium did not affect osteoclastogenesis.ConclusionInhibition of toll-like receptor 4 signaling revealed that saliva can contribute to innate immunity by preventing hematopoietic progenitors to become osteoclasts.Clinical relevanceSaliva can activate pattern recognition receptor signaling through endotoxins and other stress factors, indicating the demand for macrophages rather than for osteoclasts.
| S-EPMC5632348 | biostudies-literature
Project description:We report the results of a screen of a library of 925 potential prenyl synthase inhibitors against Trypanosoma brucei farnesyl diphosphate synthase (TbFPPS) and against T. brucei, the causative agent of human African trypanosomiasis. The most potent compounds were lipophilic analogs of the bone resorption drug zoledronate, some of which had submicromolar to low micromolar activity against bloodstream form T. brucei and selectivity indices of up to ∼ 300. We evaluated the effects of two such inhibitors on survival and parasitemia in a T. brucei mouse model of infection and found that survival increased by up to 16 days. We also investigated the binding of three lipophilic bisphosphonates to an expressed TbFPPS using crystallography and investigated the thermodynamics of binding using isothermal titration calorimetry.
| S-EPMC4649187 | biostudies-literature
Project description:Non-traditional bisphosphonates, that is, bisphosphonates that do not inhibit osteoclast viability or function, were initially reported in the 1990s by Socrates Papapoulos' group. Originally designed to study the role of the R1 residue of aminobisphosphonates on bisphosphonate affinity for hydroxyapatite, these modified bisphosphonates retained similar affinity for mineralized bone as their parent compounds, but they lacked the potential to inhibit the mevalonate pathway or bone resorption. We found that, similar to classical bisphosphonates, these non-traditional compounds prevented osteoblast and osteocyte apoptosis in vitro through a pathway that requires the expression of the gap junction protein connexin 43, and the activation of the Src/MEK/ERK signaling pathway. Furthermore, one of those compounds named IG9402 (also known as amino-olpadronate or lidadronate), was able to inhibit osteoblast and osteocyte apoptosis, without affecting osteoclast number or bone resorption in vivo in a model of glucocorticoid-induced osteoporosis. IG9402 administration also ameliorated the decrease in bone mass and in bone mechanical properties induced by glucocorticoids. Similarly, IG9402 prevented apoptosis of osteoblastic cells in a model of immobilization due to hindlimb unloading. However, in this case, the bisphosphonate was not able to preserve the bone mass, and only partially prevented the decrease in bone mechanical properties induced by immobilization. The effect of IG9402 administration was also tested in a mouse model of masticatory hypofunction through the induction of masseter muscle atrophy by unilateral injection of botulinum toxin type A (BoNTA). IG9402 partially inhibited the loss of trabecular bone microstructure in the mandibular condyle, but not the decrease in masseter muscle mass induced by BoNTA administration. In summary, these non-traditional bisphosphonates that lack anti-resorptive activity but are able to preserve osteoblast and osteocyte viability could constitute useful tools to study the consequences of preventing apoptosis of osteoblastic cells in animal models. Furthermore, they could be used to treat conditions associated with reduced bone mass and increased bone fragility in which a reduction of bone remodeling is not desirable.
| S-EPMC7138726 | biostudies-literature
Project description:Although it may sound unpleasant, camel urine has been consumed extensively for years in the Middle East as it is believed to be able to treat a wide range of diseases such as fever, cold, or even cancer. People usually take it by mixing small drops with camel milk or take it directly. The project aims to study the effects of camel urine in inhibiting the growth potential and metastatic ability of 4T1 cancer cell line in vitro and in vivo. Based on the MTT result, the cytotoxicity of camel urine against 4T1 cell was established, and it was dose-dependent. Additionally, the antimetastatic potential of camel urine was tested by running several assays such as scratch assay, migration and invasion assay, and mouse aortic ring assay with promising results in the ability of camel urine to inhibit metastatic process of the 4T1 cells. In order to fully establish camel urine's potential, an in vivo study was carried out by treating mice inoculated with 4T1 cells with 2 different doses of camel urine. By the end of the treatment period, the tumor in both treated groups had reduced in size as compared to the control group. Additional assays such as the TUNEL assay, immunophenotyping, cytokine level detection assay, clonogenic assay, and proteome profiler demonstrated the capability of camel urine to reduce and inhibit the metastatic potential of 4T1 cells in vivo. To sum up, further study of anticancer properties of camel urine is justified, as evidenced through the in vitro and in vivo studies carried out. Better results were obtained at higher concentration of camel urine used in vivo. Apart from that, this project has laid out the mechanisms employed by the substance to inhibit the growth and the metastatic process of the 4T1 cell.
| S-EPMC5739131 | biostudies-literature
Project description:Estrogens have been implicated as complete carcinogens for breast and other tissues through mechanisms involving increased cell proliferation, oxidative stress, and DNA damage. Because of their potent antioxidant activity and other effects, tocopherols have been shown to exert antitumor activities in various cancers. However, limited information is available on the effect of different forms of tocopherols in estrogen-mediated breast cancer. To address this, we examined the effects of α-, γ-, and δ-tocopherols as well as a natural γ-tocopherol-rich mixture of tocopherols, γ-TmT, on estrogen-stimulated MCF-7 cells in vitro and in vivo For the in vivo studies, MCF-7 cells were injected into the mammary fat pad of immunodeficient mice previously implanted with estrogen pellets. Mice were then administered diets containing 0.2% α-, γ-, δ-tocopherol, or γ-TmT for 5 weeks. Treatment with α-, γ-, δ-tocopherols, and γ-TmT reduced tumor volumes by 29% (P < 0.05), 45% (P < 0.05), 41% (P < 0.05), and 58% (P < 0.01), as well as tumor weights by 20%, 37% (P < 0.05), 39% (P < 0.05), and 52% (P < 0.05), respectively. γ- and δ-tocopherols and γ-TmT inhibited the expression of cell proliferation-related genes such as cyclin D1 and c-Myc, and estrogen-related genes such as TFF/pS2, cathepsin D, and progesterone receptor in estrogen-stimulated MCF-7 cells in vitro Further, γ- and δ-tocopherols decreased the levels of estrogen-induced oxidative stress and nitrosative stress markers, 8-hydroxy-2'-deoxyguanosine and nitrotyrosine, as well as the DNA damage marker, γ-H2AX. Our results suggest that γ- and δ-tocopherols and the γ-tocopherol-rich mixture are effective natural agents for the prevention and treatment of estrogen-mediated breast cancer. Cancer Prev Res; 10(3); 188-97. ©2017 AACR.
| S-EPMC5337152 | biostudies-literature
Project description:BackgroundCell cycle regulatory pathway is a well-established pathway mainly dependent on cyclin-dependent kinases (CDKs), which are regulated positively by cyclins and negatively by cyclin-dependent kinase inhibitors(CKIs). Cyclin-dependent kinase 2 associate protein 1(CDK2-AP1) is a specific negative regulatory protein for CDK2, is important in the cancer cell cycle. However, the function of CDK2-AP1 in breast cancer remains unclear. We designed therefore explored the effects of CDK2-AP1 on breast cancer growth and its chemo-sensitivity.MethodsExpression of CDK2-AP1, CDK2 and CyclinD1 in 209 cases of pathological specimens using IHC staining was measured. Lost-of-function and Gain-of-function assays were used in vivo and in vitro relating to the specific role of CDK2-AP1 in breast cancer. We analyzed in vivo and in vitro the impact of CDK2-AP1 on chemotherapy sensitivity in breast cancer.ResultsThe positive ratio of CDK2-AP1 expression was reduced successively in normal breast tissue, DCIS, invasive breast cancer and relapsed breast cancer, however, with CDK2 and CyclinD1 it was suggested that CDK2-AP1 was correlated closely with the tumorigenesis and progress, and might work as a tumor suppressor. After down-regulating CDK2-AP1 in breast cancer cells, the cell cycle was accelerated and cell proliferation enhanced. The cell cycle was arrested in G0/G1 phase and G2/M phase after up-regulating CDK2-AP1 in breast cancer cells, inhibiting cell proliferation. The expression of CDK2 and CyclinD1 changed accordingly after downregulation or upregulation of CDK2-AP1 by western blot, suggesting a role of the CDK2-AP1/CDK2/CyclinD1 cell cycle pathway in the initiation and progression of breast cancer. Similar results were obtained in animal assays. The data indicates that CDK2-AP1 can induce sensitivity to docetaxel treatment in breast cancer cells.ConclusionsCDK2-AP1 affects tumorigenesis, tumor growth and chemo-sensitivity by cell cycle regulation, which can potentially to be a therapeutical agent in breast cancer.
| S-EPMC4279590 | biostudies-literature
Project description:Adipose tissue expansion is accompanied by infiltration and accumulation of pro-inflammatory macrophages, which links obesity to pathologic conditions such as type 2 diabetes. However, little is known regarding the role of pro-inflammatory adipose tissue remodeling in the thermogenic activation of brown/beige fat. Here, we investigated the effect of pattern recognition receptors (PRR) activation in macrophages, especially the toll-like receptor 4 (TLR4) and Nod-like receptor 3 (NLRP3), on white adipocyte browning. We report that TLR4 activation by lipopolysaccharide repressed white adipocyte browning in response to ?3-adrenergic receptor activation and caused ROS production and mitochondrial dysfunction, while genetic deletion of TLR4 protected mitochondrial function and thermogenesis. In addition, activation of NLRP3 inflammasome in macrophages attenuated UCP1 induction and mitochondrial respiration in cultures of primary adipocytes, while the absence of NLRP3 protected UCP1 in adipocytes. The effect of NLRP3 inflammasome activation on browning was mediated by IL-1? signaling, as blocking IL-1 receptor in adipocytes protected thermogenesis. We also report that IL-1? interferes with thermogenesis via oxidative stress stimulation and mitochondrial dysfunction as we observed a statistically significant increase in ROS production, decrease in SOD enzyme activity, and increase in mitochondrial depolarization in adipocytes treated with IL-1?. Collectively, we demonstrated that inflammatory response to obesity, such as TLR4 and NLRP3 inflammasome activation as well as IL-1? secretion, attenuates ?3-adrenoreceptor-induced beige adipocyte formation via oxidative stress and mitochondrial dysfunction. Our findings provide insights into targeting innate inflammatory system for enhancement of the adaptive thermogenesis against obesity.
| S-EPMC6066287 | biostudies-literature
Project description:Background: Anlotinib is a highly potent multi-target tyrosine kinase inhibitor. Accumulating evidence suggests that anlotinib exhibits effective anti-tumor activity against various cancer subtypes. However, the effects of anlotinib against cisplatin-resistant (CIS) ovarian cancer (OC) are yet to be elucidated. The objective of this study was to investigate the inhibitory effect of anlotinib on the pathogenesis of cisplatin-resistant OC. Materials and Methods: Human OC cell lines (A2780 and A2780 CIS) were cultured and treated with or without anlotinib. The effects of anlotinib on cell proliferation were determined using cell-counting kit-8 and colony-formation assays. To evaluate the invasion and metastasis of OC cells, we performed wound-healing and transwell assays. The cell cycle was analyzed via flow cytometry. A xenograft mouse model was used to conduct in vivo studies to verify the effects of anlotinib. The expression of Ki-67 in the tumor tissue was detected via immunohistochemistry. Quantitative real-time polymerase chain reaction and Western blotting were used to measure the mRNA and protein levels. Results: Our study revealed that anlotinib significantly inhibited the proliferation, migration, and invasion of A2780 and A2780 CIS in a dose-dependent way in vitro (p < 0.05). Through R software ‘limma’ package analysis of GSE15372, it was found that, in comparison with A2780, PLK2 was expressed in significantly low levels in the corresponding cisplatin-resistant strains. The ERK1/2/Plk2 signaling axis mediates the inhibitory effect of anlotinib on the proliferation and migration of ovarian cancer cell lines. Moreover, our research found that anlotinib effectively inhibited the growth of tumor cells in an OC xenograft mouse model. Conclusions: In this study, anlotinib showed excellent inhibitory effects against cisplatin-resistant OC both in vitro and in vivo. These results add to the growing body of evidence supporting anlotinib as a potential anticancer agent against OC.
| S-EPMC9785832 | biostudies-literature
Project description:The human specific bacterial pathogen Helicobacter pylori (H. pylori) is associated with severe gastric diseases, including gastric cancer. Recently, the increasing resistance makes the usage of antibiotics less effectively. Therefore, development of a new antimicrobial agent is required to control H. pylori infection. In the current study, the inhibitory effect of β-caryophyllene on H. pylori growth, as well as the antibacterial therapeutic effect, has been demonstrated. β-caryophyllene inhibited H. pylori growth via the downregulation of dnaE, dnaN, holB, and gyrA and also downregulated virulence factors such as CagA, VacA, and SecA proteins. β-caryophyllene inhibited expression of several T4SS components, so that CagA translocation into H. pylori-infected AGS gastric cancer cells was decreased by β-caryophyllene treatment. β-caryophyllene also inhibited VacA entry through the downregulation of T5aSS. After β-caryophyllene administration on Mongolian gerbils, the immunohistochemistry (IHC) and Hematoxylin&Eosin stains showed therapeutic effects in the treated groups. Hematological data, which was consistent with histological data, support the therapeutic effect of β-caryophyllene administration. Such a positive effect of β-caryophyllene on H. pylori infection potently substantiates the natural compound as being capable of being used as a new antimicrobial agent or functional health food to help patients who are suffering from gastroduodenal diseases due to H. pylori infection.
| S-EPMC7037973 | biostudies-literature