Mammary Cells Research Articles

Hg(II)/Fe(III) Turn‐On Fluorescence Response of Rhodamine‐Anthracene Derivatives with Variable Diamine Chain Length

AbstractTwo rhodamine‐anthracene derivative probes (RA1 and RA2) were designed and synthesized to incorporate different lengths of diamine chains between the rhodamine and anthracene moieties. Among the 16 different metal ions tested, RA1 exhibited the highest fluorescence enhancement in response to Hg2+ with a limit of detection (LOD) of 3.76×10−7 M, while RA2 showed the greatest fluorescence enhancement in the presence of Fe3+ with the LOD of 2.41×10−7 M. Notably, RA1 effectively eliminated the interference caused by Fe3+ during Hg2+ detection, whereas RA2 successfully eliminated the interference of Hg2+ during Fe3+ determination. This response difference between the two ions holds particular importance in scenarios where both heavy metal ions coexist. Additionally, fluorescence microscopy imaging of mouse mammary cancer cells (4T1) under the influence of Fe3+ demonstrated the potential applications of the probe. This study highlights the significance of a simple molecular design approach in tailoring probe selectivity by modifying the intramolecular flexible chain length.

Escherichia coli infection mediates pyroptosis via activating p53-p21 pathway-regulated apoptosis and cell cycle arrest in bovine mammary epithelial cells

Escherichia coli (E. coli) is a major environmental pathogen that causes mammary tissue damage and cell death, which results in substantial economic losses. Pyroptosis, a novel form of programmed cell death characterized by DNA fragmentation, chromatin condensation, cell swelling and leakage of cell contents, often occurs after inflammatory apoptotic pathways activation. Our objective was to investigate the intraction between E. coli infection and bovine mammary epithelial cells (bMECs) with pyroptosis and to explore the underlying regulatory mechanism. bMECs were infected with E. coli for 6 h. Lactic dehydrogenase activities, interleukin (IL)-10, IL-1β, IL-18 and tumor necrosis factor-α concentrations, total apoptosis indexes, and protein expressions of P-cdc25c, P-CDK1, cleaved caspase 9, cleaved caspase 3, cleaved PARP, P–NF-κB, NLRP3, ASC, caspase 1, gasdermin D N-terminal, IL-1β and IL-18 were significantly increased in E. coli infected bMECs. Whereas, cell membrane potential, protein levels of cdc25c, CDK1, cyclin B1, and Bcl-2/Bax level were markedly reduced. Furthermore, Ac-DEVD-CHO (specific inhibitor of apoptosis) dramatically suppressed pyroptosis in bMECs. Moreover, expressions of p53 and p21 promptly improved after E. coli infection, however, Pifithrin-α (specific inhibitor of p53) inhibited p53-p21 pathway, apoptosis, cell cycle arrest and pyroptosis. These results elaborated that E. coli infection of bMECs induced pyroptosis through activating the p53-p21 pathway-mediated apoptosis and cell cycle arrest. Taken together, inhibition of pyroptosis via suppressing of p53-p21 pathway may be an effective therapeutic approach for treating E. coli-induced mastitis, offering efficient theoretical support for the protection and treatment of bovine mastitis.

Identification of the SNARE complex that mediates the fusion of multivesicular bodies with the plasma membrane in exosome secretion.

Exosomes play crucial roles in local and distant cellular communication and are involved in various physiological and pathological processes. Tumour-derived exosomes are pivotal to tumorigenesis, but the precise mechanisms underlying their secretion remain elusive. In particular, the SNARE proteins that mediate the fusion of multivesicular bodies (MVBs) with the plasma membrane (PM) in tumour cells are subject to debate. In this study, we identified syntaxin-4, SNAP-23, and VAMP-7 as the SNAREs responsible for exosome secretion in MCF-7 breast cancer cells and found that a SNARE complex consisting of these SNAREs can drive membrane fusion in vitro. Deletion of any of these SNAREs in MCF-7 cells did not affect MVB biogenesis and transportation, indicating their specific involvement in MVB-PM fusion. In addition, syntaxin-4, SNAP-23, and VAMP-7 play equivalent roles in exosome secretion in both HeLa cervical cancer cells and A375 melanoma cells, suggesting their conserved function in exosome secretion. Furthermore, deletion of VAMP-7 in 4T1 mammary carcinoma cells efficiently inhibited exosome secretion and led to significant attenuation of tumour growth and lung metastasis in mouse models, implying that VAMP-7 may hold promise as a novel therapeutic target for breast cancer.

Antiproliferative effect of methanolic extract of Tamarindus indica in C127I cell line

Investigation of natural bioactive compounds that possess antioxidant and anticancer properties have drawn interest in scientific and industrial perspectives. The current study was conducted to assess the antiproliferative action of methanolic extract of seed coat of Tamarindus indica in C127I mouse mammary tumour cell line. 3-(4,5-dimethyl thazol-2-yl)-2, 5-diphenyl tetrazolium bromide (MTT) assay is performed to assess the cytotoxicity of methanolic extracts of T. indica in the C127I cell line at concentrations of 320, 160, 80, 40, 20, and 10 µg/mL. The half maximal inhibitory concentration (IC50) was then calculated using Graph Pad Prism 5.0. Doxorubicin was used as positive control. After determining IC50, the cells were treated with extract of T. indica at half, IC50 and double IC50 concentrations after being seeded at a concentration of 1x105 cells/mL in 6 well plates. After 24 hours the cells were trypsinised and stained with acridine orange-ethidium bromide (AOEB) to assess the morphology of apoptosis. When the cells were exposed to various doses of the extract, a dose-dependent decrease in cell viability was observed, and the IC50 value of T. indica was estimated to be 13 µg/ml. In the control cells, AO/EB labelling revealed proliferating cells with green fluorescence, but the T. indica extract-treated cells displayed dose dependent shift from orange to red fluorescence. In conclusion, methanolic extract of Tamarindus indica was found to possess antiproliferative effect in C127I cell line comparable to doxorubicin.

Compaction of Calf Thymus DNA by a Potential One-Head-Two-Tail Surfactant: Properties of Nanomaterials and Biological Testing for Gene Delivery.

A one-head-two-tail cationic surfactant, Dilauryldimethylammonium bromide (DDAB) has shown a great extent of calf thymus DNA (ct-DNA) compaction being adsorbed on the surfaces of negatively charged SiO2 nanoparticles (NPs). DDAB molecules show high adsorption efficiency and induce many positive surface charges per-unit surface area of the SiO2 NPs compared to cationic Gemini (12-6-12) and conventional (DTAB) surfactants in an aqueous medium at pH 7.4, as evident from zeta potential and EDAX data. Transmission electron microscopy and field emission scanning electron microscopy images, along with ethidium bromide exclusion assay and DLS data support the compaction of ct-DNA. Fluorescence microscopic images show that in the presence of SiO2 NPs, DDAB can perform 50% compaction of ct-DNA at a concentration ∼58% and ∼99% lower than that of 12-6-12 and DTAB, respectively. Better ct-DNA compaction by DDAB is evident compared to other Gemini surfactants (12-4-12 and 12-8-12) as well reported before. Time-correlated single photon counting fluorescence intensity decay measurements of a probe DAPI in ct-DNA have revealed the average lifetime value that is decreased by ∼61% at 2.5 μM of DDAB in the presence of SiO2 NPs as compared to a decrease by only ∼29% in its absence, supporting NPs-induced stronger surfactant binding with ct-DNA. Fluorescence lifetime data have also demonstrated the crowding effect of NPs. At 2.5 μM of DDAB, both fast and slow rotational relaxation components of DAPI contribute almost equally to depolarization with the absence of NPs; however, with the presence of NPs, ∼96% weightage of the anisotropy decay is for the fast component. The present DDAB-SiO2 NPs combination has proved to be an excellent gene delivery system based on the cell viability in the mouse mammary gland adenocarcinoma cells (4T1) and human embryonic kidney (HEK) 293 cell lines, and in vitro and in vivo studies.

Schizophrenia plausible protective effect of microRNA-137 is potentially related to estrogen and prolactin in female patients.

Schizophrenia (SCZ) is a serious chronic mental disorder. Our previous case-control genetic association study has shown that microRNA-137 (miR-137) may only protect females against SCZ. Since estrogen, an important female sex hormone, exerts neuroprotective effects, the relationship between estrogen and miR-137 in the pathophysiology of SCZ was further studied in this study. Genotyping of single-nucleotide polymorphism rs1625579 of miR-137 gene in 1,004 SCZ patients and 896 healthy controls was conducted using the iMLDR assay. The effect of estradiol (E2) on the miR-137 expression was evaluated on the human mammary adenocarcinoma cell line (MCF-7) and the mouse hippocampal neuron cell line (HT22). The relationships between serum E2, prolactin (PRL), and peripheral blood miR-137 were investigated in 41 SCZ patients and 43 healthy controls. The miR-137 and other reference miRNAs were detected by real-time fluorescent quantitative reverse transcription-PCR. Based on the well-known SNP rs1625579, the distributions of protective genotypes and alleles of the miR-137 gene were not different between patients and healthy controls but were marginally significantly lower in female patients. E2 upregulated the expression of miR-137 to 2.83 and 1.81 times in MCF-7 and HT22 cells, respectively. Both serum E2 and blood miR-137 were significantly decreased or downregulated in SCZ patients, but they lacked expected positive correlations with each other in both patients and controls. When stratified by sex, blood miR-137 was negatively correlated with serum E2 in female patients. On the other hand, serum PRL was significantly increased in SCZ patients, and the female patients had the highest serum PRL level and a negative correlation between serum PRL and blood miR-137. The plausible SCZ-protective effect of miR-137 may be female specific, of which the underlying mechanism may be that E2 upregulates the expression of miR-137. This protective mechanism may also be abrogated by elevated PRL in female patients. These preliminary findings suggest a new genetic/environmental interaction mechanism for E2/miR-137 to protect normal females against SCZ and a novel E2/PRL/miR-137-related pathophysiology of female SCZ, implying some new antipsychotic ways for female patients in future.

Immunoregulatory and Anti-cancer Activities of Combination Treatment of Novel Four-Herb Formula and Doxorubicin in 4T1-Breast Cancer Bearing Mice.

To investigate the in vivo immunomodulatory and anti-tumor mechanisms of the combined treatment of novel Four-Herb formula (4HF) and doxorubicin in triple-negative breast cancer (TNBC). Murine-derived triple-negative mammary carcinoma cell line, 4T1 cells, was cultured and inoculated into mouse mammary glands. Sixty-six mice were randomly assigned into 6 groups (n=11 in ench): naïve, control, LD 4HF (low dose 4HF), HD 4HF (high dose 4HF), LD 4HF + D (low dose and doxorubicin), and D (doxorubicin). Apart from the naïve group, each mouse received subcutaneous inoculation with 5 × 105 4T1 cells resuspended in 100 µL of normal saline in the mammary fat pads. Starting from the day of tumor cell inoculation, tumors were grown for 6 days. The LD and HD groups received daily oral gavage of 658 and 2,630 mg/kg 4HF, respectively. The LD 4HF+D group received daily oral gavage of 658 mg/kg 4HF and weekly intraperitoneal injection of doxorubicin (5 mg/kg). The D group received weekly intraperitoneal injections of doxorubicin (5 mg/kg). The treatment naïve mice received daily oral gavage of 0.2 mL double distilled water and 0.1 mL normal saline via intraperitoneal injection once a week. The control group received daily oral gavage of 0.2 mL double-distilled water. The treatment period was 30 days. At the end of treatment, mice organs were harvested to analyze immunological activities via immunophenotyping, gene and multiplex analysis, histological staining, and gut microbiota analysis. Mice treated with the combination of 4HF and doxorubicin resulted in significantly reduced tumor and spleen burdens (P<0.05), altered the hypoxia and overall immune lymphocyte landscape, and manipulated gut microbiota to favor the anti-tumor immunological activities. Moreover, immunosuppressive genes, cytokines, and chemokines such as C-C motif chemokine 2 and interleukin-10 of tumors were significantly downregulated (P<0.05). 4HF-doxorubicin combination treatment demonstrated synergetic activities and was most effective in activating the anti-tumor immune response (P<0.05). The above results provide evidence for evaluating the immune regulating mechanisms of 4HF in breast cancer and support its clinical significance in its potential as an adjunctive therapeutic agent or immune supplement.

Sirtuin 1 Expression in Canine Mammary Tumors: A Pilot Study.

Sirtuin 1 (SIRT1) is a protein involved in aging, cell protection, and energy metabolism in mammals. Recently, SIRT1 has been intensively studied in medical oncology, but the role of SIRT1 is still controversial, as it has been proposed as both an oncogene and a tumor suppressor. The aim of this study is to investigate the expression of SIRT1 by immunohistochemistry in canine mammary tissues, and by Western blot and immunofluorescence analysis in different canine mammary cell lines. Our results showed a decrease in SIRT1 expression from normal mammary gland tissue, and from benign and well-differentiated malignant tumors (G1) to less differentiated ones (G2-G3). Furthermore, a shift in the subcellular localization of SIRT1 from the nucleus to the cytoplasm was observed in less differentiated malignant tumors. However, further studies are needed to investigate the subcellular localization of SIRT1 in canine cancer cells and the role it may play in oncogenesis in animals.

MiR-203 drives breast cancer cell differentiation

A hallmark of many malignant tumors is dedifferentiated (immature) cells bearing slight or no resemblance to the normal cells from which the cancer originated. Tumor dedifferentiated cells exhibit a higher capacity to survive to chemo and radiotherapies and have the ability to incite tumor relapse. Inducing cancer cell differentiation would abolish their self-renewal and invasive capacity and could be combined with the current standard of care, especially in poorly differentiated and aggressive tumors (with worst prognosis). However, differentiation therapy is still in its early stages and the intrinsic complexity of solid tumor heterogeneity demands innovative approaches in order to be efficiently translated into the clinic. We demonstrate here that microRNA 203, a potent driver of differentiation in pluripotent stem cells (ESCs and iPSCs), promotes the differentiation of mammary gland tumor cells. Combining mouse in vivo approaches and both mouse and human-derived tridimensional organoid cultures, we report that miR-203 influences the self-renewal capacity, plasticity and differentiation potential of breast cancer cells and prevents tumor cell growth in vivo. Our work sheds light on differentiation-based antitumor therapies and offers miR-203 as a promising tool for directly confronting the tumor-maintaining and regeneration capability of cancer cells.

Irradiated Mammary Spheroids Elucidate Mechanisms of Macrophage-Mediated Breast Cancer Recurrence.

While most patients with triple negative breast cancer receive radiation therapy to improve outcomes, a significant subset of patients continue to experience recurrence. Macrophage infiltration into radiation-damaged sites has been shown to promote breast cancer recurrence in pre-clinical models. However, the mechanisms that drive recurrence are unknown. Here, we developed a novel spheroid model to evaluate macrophage-mediated tumor cell recruitment. We characterized infiltrating macrophage phenotypes into irradiated mouse mammary tissue via flow cytometry. We then engineered a spheroid model of radiation damage with primary fibroblasts, macrophages, and 4T1 mouse mammary carcinoma cells using in vivo macrophage infiltration results to inform our model. We analyzed 4T1 infiltration into spheroids when co-cultured with biologically relevant ratios of pro-healing M2:pro-inflammatory M1 macrophages. Finally, we quantified interleukin 6 (IL-6) secretion associated with conditions favorable to tumor cell infiltration, and we directly evaluated the impact of IL-6 on tumor cell invasiveness in vitro and in vivo. In our in vivo model, we observed a significant increase in M2 macrophages in mouse mammary glands 10days post-irradiation. We determined that tumor cell motility toward irradiated spheroids was enhanced in the presence of a 2:1 ratio of M2:M1 macrophages. We also measured a significant increase in IL-6 secretion after irradiation both in vivo and in our model. This secretion increased tumor cell invasiveness, and tumor cell invasion and recruitment were mitigated by neutralizing IL-6. Our work suggests that interactions between infiltrating macrophages and damaged stromal cells facilitate breast cancer recurrence through IL-6 signaling. The online version contains supplementary material available at 10.1007/s12195-023-00775-x.

Radical Tumor Denervation Activates Potent Local and Global Cancer Treatment.

This preliminary study seeks to determine the effect of R&P denervation on tumor growth and survival in immunocompetent rats bearing an aggressive and metastatic breast solid tumor. A novel microsurgical approach was applied "in situ", aiming to induce R&P denervation through the division of every single nerve fiber connecting the host with the primary tumor via its complete detachment and re-attachment, by resecting and reconnecting its supplying artery and vein (anastomosis). This preparation, known as microsurgical graft or flap, is radically denervated by definition, but also effectively delays or even impedes the return of innervation for a significant period of time, thus creating a critical and therapeutic time window. Mammary adenocarcinoma cells (HH-16.cl4) were injected into immunocompetent Sprague Dawley adult rats. When the tumors reached a certain volume, the subjects entered the study. The primary tumor, including a substantial amount of peritumoral tissue, was surgically isolated on a dominant artery and vein, which was resected and reconnected using a surgical microscope (orthotopic tumor auto-transplantation). Intending to simulate metastasis, two or three tumors were simultaneously implanted and only one was treated, using the surgical technique described herein. Primary tumor regression was observed in all of the microsurgically treated subjects, associated with a potent systemic anticancer effect and prolonged survival. In stark contrast, the subjects received a close to identical surgical operation; however, with the intact neurovascular connection, they did not achieve the therapeutic result. Animals bearing multiple tumors and receiving the same treatment in only one tumor exhibited regression in both the "primary" and remote- untreated tumors at a clinically significant percentage, with regression occurring in more than half of the treated subjects. A novel therapeutic approach is presented, which induces the permanent regression of primary and, notably, remote tumors, as well as, evidently, the naturally occurring metastatic lesions, at a high rate. This strategy is aligned with the impetus that comes from the current translational research data, focusing on the abrogation of the neuro-tumoral interaction as an alternative treatment strategy. More data regarding the clinical significance of this are expected to come up from a pilot clinical trial that is ongoing.

Unveiling the mechanisms underlying photothermal efficiency of gold shell-isolated nanoparticles (AuSHINs) on ductal mammary carcinoma cells (BT-474)

Gold nanoparticles are valuable photothermal agents owing to their efficient photothermal conversion, photobleaching resistance, and potential surface functionalization. Herein, we combined bioinspired membranes with in vitro assays to elicit the molecular mechanisms of gold shell-isolated nanoparticles (AuSHINs) on ductal mammary carcinoma cells (BT-474). Langmuir and Langmuir-Schaefer (LS) films were handled to build biomembranes from BT-474 lipid extract. AuSHINs incorporation led to surface pressure-area (π-A) isotherms expansion, increasing membrane flexibility. Fourier-transform infrared spectroscopy (FTIR) of LS multilayers revealed electrostatic AuSHINs interaction with head portions of BT-474 lipid extract, causing lipid chain disorganization. Limited AuSHINs insertion into monolayer contributed to hydroperoxidation of the unsaturated lipids upon irradiation, consistently with the surface area increments of ca. 2.0%. In fact, membrane disruption of irradiated BT-474 cells containing AuSHINs was confirmed by confocal microscopy and LDH leakage, with greater damage at 2.2 × 1013 AuSHINs/mL. Furthermore, the decrease in nuclei dimensions indicates cell death through photoinduced damage.

Cancer Cell Membrane Wrapped Nanoparticles for the Delivery of a Bcl-2 Inhibitor to Triple-Negative Breast Cancer.

Overexpression of the antiapoptotic protein B-cell lymphoma 2 (Bcl-2) is correlated with poor survival outcomes in triple-negative breast cancer (TNBC), making Bcl-2 inhibition a promising strategy to treat this aggressive disease. Unfortunately, Bcl-2 inhibitors developed to date have limited clinical success against solid tumors, owing to poor bioavailability, insufficient tumor delivery, and off-target toxicity. To circumvent these problems, we loaded the Bcl-2 inhibitor ABT-737 in poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) that were wrapped with phospholipid membranes derived from 4T1 murine mammary cancer cells, which mimic the growth and metastasis of human TNBC. We show that the biomimetic cancer cell membrane coating enabled the NPs to preferentially target 4T1 TNBC cells over noncancerous mammary epithelial cells in vitro and significantly increased NP accumulation in orthotopic 4T1 tumors in mice after intravenous injection by over 2-fold compared to poly(ethylene glycol)-poly(lactide-co-glycolic) (PEG-PLGA) copolymer NPs. Congruently, the ABT-737 loaded, cancer cell membrane-wrapped PLGA NPs (ABT CCNPs) induced higher levels of apoptosis in TNBC cells in vitro than ABT-737 delivered freely or in PEG-PLGA NPs. When tested in a syngeneic spontaneous metastasis model, the ABT CCNPs significantly increased apoptosis (evidenced by elevated active caspase-3 and decreased Bcl-2 staining) and decreased proliferation (denoted by reduced Ki67 staining) throughout tumors compared with saline or ABT-loaded PEG-PLGA NP controls. Moreover, the ABT CCNPs did not alter animal weight or blood composition, suggesting that the specificity afforded by the TNBC cell membrane coating mitigated the off-target adverse effects typically associated with ABT-737. Despite these promising results, the low dose of ABT CCNPs administered only modestly reduced primary tumor growth and metastatic nodule formation in the lungs relative to controls. We posit that increasing the dose of ABT CCNPs, altering the treatment schedule, or encapsulating a more potent Bcl-2 inhibitor may yield more robust effects on tumor growth and metastasis. With further development, drug-loaded biomimetic NPs may safely treat solid tumors such as TNBC that are characterized by Bcl-2 overexpression.

Extracellular vesicles in the treatment and diagnosis of breast cancer: a status update.

Breast cancer is one of the leading causes of cancer-related death in women. Currently, the treatment of breast cancer is limited by the lack of effectively targeted therapy and patients often suffer from higher severity, metastasis, and resistance. Extracellular vesicles (EVs) consist of lipid bilayers that encapsulate a complex cargo, including proteins, nucleic acids, and metabolites. These bioactive cargoes have been found to play crucial roles in breast cancer initiation and progression. Moreover, EV cargoes play pivotal roles in converting mammary cells to carcinogenic cells and metastatic foci by extensively inducing proliferation, angiogenesis, pre-metastatic niche formation, migration, and chemoresistance. The present update review mainly discusses EVs cargoes released from breast cancer cells and tumor-derived EVs in the breast cancer microenvironment, focusing on proliferation, metastasis, chemoresistance, and their clinical potential as effective biomarkers.

Proliferative response of ERα-positive breast cancer cells to 10 μM enterolactone, and the associated alteration in the transcriptomic landscape

Enterolactone (EL) is a product of gut-microbial metabolism of dietary plant lignans. Studies linking EL with breast cancer risk have bolstered investigations into its effects on the mammary epithelial cells, and the mechanisms thereof. While it binds to the estrogen receptor α (ERα), its effect on the proliferation of mammary tumor cell lines is reportedly ambivalent; depending on its concentration. The genomic correlates of EL actions also remain unexplored. Here we have elaborately studied the effect of EL on proliferation of ERα-positive, and ERα-negative cell lines. 10 µM EL significantly enhanced the growth of the ERα-positive MCF-7 or T47D breast cancer cells, but not the ERα-negative MDA-MB-231 or MDA-MB-453 cells. In MCF-7 cells, it significantly increased the expression of TFF1 mRNA, an estrogen-induced transcript. The binding of ERα to the estrogen response element within the TFF1 locus further demonstrated the pro-estrogenic effect of 10 µM EL. We further explored the genome-wide transcriptomic effect of 10 µM EL using the next generation sequencing technology (RNA-seq). Analysis of RNA-seq data obtained from vehicle (0.1% DMSO)- or 10 µM EL-treated MCF-7 cells revealed modulation of expression of diverse sets of functionally related genes, which reflected cell cycle progression. The manner in which 10 µM EL regulated the hallmark G2/M checkpoint, and estrogen-response-late genes correlated with proliferation inducing, and estrogen-like effects of EL on MCF-7 cells.

Synthesis and Cytotoxicity Evaluation of Tyrosine-5 Fluorinated Analogues of RA-VII, An Antitumor Bicyclic Hexapeptide

AbstractRA-VII analogues fluorinated at each ε-position of Tyr-5 were designed. The synthesis of these peptide analogues commenced with the preparation of atropisomeric fluorocycloisodityrosines from protected 3-fluoro-L-tyrosyl-3-boronyl-L-tyrosine mediated by copper(II) acetate and 4-(dimethylamino)pyridine. After N-methylation, the tetrapeptide segment was coupled with the N-terminus of each fluorocycloisodityrosine to afford a hexapeptide. After removal of the C- and N-terminal protecting groups, each peptide was subjected to macrocyclization to produce an analogue. The analogue with a β-oriented fluorine atom was equipotent to RA-VII with regard to cytotoxicity toward human mammary carcinoma MCF-7 cells, and showed 2.1-fold and 1.4-fold lower cytotoxicities than RA-VII toward human promyelocytic leukemia HL-60 and human colorectal cancer HCT-116 cells, respectively, whereas the analogue with an α-oriented fluorine atom showed 7.7-fold, 6.0-fold, and 14-fold lower cytotoxicities than RA-VII toward those cells, respectively.

Evaluation of in vitro immunostimulatory and cytotoxic effects of recombinant survivin protein in combination with doxorubicin and breast cancer antigen-loaded polycaprolactone nanoparticles

In this study, the immunostimulatory and anticancer activities of the doxorubicin (DOX), MCF-7 and MDA-MB-231 breast cancer antigen-loaded polycaprolactone (PCL) nanoparticles (NPs) in combination with survivin recombinant protein (RP) and an alum adjuvant are evaluated in vitro on J774 macrophage, MCF-7 and MDA-MB-231 breast cancer cell lines. A double-emulsion solvent evaporation method was used for encapsulation of DOX and antigens into PCL NPs. The physicochemical characterization of NPs included size, morphology, zeta potential, release profiles and encapsulation efficiencies, analyzed using scanning electron microscopy, a zeta-sizer and UV–vis spectrometry. The cytotoxic and inhibitory effects of NPs were determined using a methyl thiazolyl tetrazolium assay. Immunostimulatory effects of the NPs were detected by Griess reaction and ELISA tests to determine nitric oxide and cytokine levels, respectively. According to the results, DOX and antigen-loaded PCL NPs ranged between 240 nm and 290 nm. Antigen and drug-loaded NPs appear less toxic over macrophage cells in comparison with non-capsulated free agents. In addition, considerable inhibitory effects of antigen and drug-loaded NPs were observed at non-toxic concentrations, such as 25 and 50 μg ml−1, on human mammary cancer cell lines (p⩽ 0.0001). The amount of nitrite released from macrophages that were treated with antigen and DOX-encapsulated PCL NPs, in combination with alum and survivin RP, after 96 h incubation was significantly higher than the control, especially at 50 and 100 μg ml−1, and triggered macrophages to produce high quantities of IL-4 and IL-12 cytokines in contrast to the control. As a result, DOX and antigen-loaded PCL NPs in combination with survivin and alum adjuvant revealed significant immunostimulatory and inhibiting influence on macrophage and breast cancer cells, respectively. The outcomes revealed that antigen and drug-loaded PCL NPs supplemented with survivin RP and an alum adjuvant created an effective platform for the development of nanotechnology-based immunotherapeutic tools to inhibit breast cancer cells. However, these outputs should be supported by further in vivo studies.

Inversion of a paradigm: The positive roles of plant phenolics in dairy goat nutrition

The paradigm prevailing in the nineties, based on studies with goats fed tannin-rich browse as sole feed, was that dietary tannins are associated with impaired feed intake, increased fecal loss of N, and impaired NDF digestibility, hence, impaired productivity, with much research aiming at alleviating these negative effects. However, dairy goats fed plant phenolic-rich diets, and supplemented with concentrates will voluntarily ingest diets comprising more than 4% of tannins, without impairment of total feed intake and milk yield. Also, supplementing them with Poly (ethylene-glycol) MW 4000 is not needed. All plant phenolics, including condensed tannins, are partly digested, inferring that phenolic derivatives are absorbed and may reach tissues. In some studies, diets rich in plant phenolics are associated with the production of milk richer in protein and fat, reduced urea content, and improved curdling performance. Feeding goats with plant phenolics-rich diets is generally associated with inhibited bio-hydrogenation (BH) of fatty acids (FA) in the rumen, resulting in decreased saturated FA, increased polyunsaturated FA (PUFA) and, in particular C18:3–n3 but also a modified composition of de novo synthesized FA in goat milk that suggest direct action of phenolics on the mammary gland. Knowledge of the absorption of plant phenolics is still scarce for goats but even modest daily intakes seem to be associated with increased phenolic derivatives and anti-oxidative activity in plasma and milk. Dietary phenolic compounds may induce a local effect in the mammary gland and its production units (the mammary epithelium cells) encompassing increased cellular triglyceride and casein contents, and increased ATP production and non-mitochondrial oxygen consumption. Plant phenolics and their derivatives divert energy to production of milk fat, protein and lactose, with less energy directed to cellular damage control. Dietary plant phenolics are associated with improved goat health and production of healthier milk. In addition, dietary plant phenolics have anti-parasitic properties and are environment-friendly as they decrease gut methane emission and they increase fecal N at the expense of rapidly decomposable urinary N. Therefore, the attitude of scientists towards feeding plant phenolics has evoluted from mostly negative to mostly positive.

Vanillic acid exhibits potent antiproliferative and free radical scavenging effects under in vitro conditions

Cells endure oxidative stress as a result of an imbalance between the status of body’s reactive oxygen species (ROS) and antioxidants. Higher production of ROS or weak antioxidant defense mechanism in the cell could lead to various pathological disorders, including carcinogenesis. The present study investigated the in vitro free radical scavenging activity and antiproliferative efficacy of vanillic acid using a spectrum of radical scavenging assays and cytotoxic assays, respectively. Vanillic acid’s antioxidant ability was investigated using in vitro antioxidant assays such as 2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), hydroxyl, and superoxide radical scavenging assays. The current study observed an excellent free radical scavenging capacity of vanillic acid, which was comparable to that of ascorbic acid (reference drug). The antiproliferative effect of vanillic acid was assessed in mammary cancer cells (Michigan Cancer Foundation-7 [MCF-7]) by using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, ROS generation potential, changes in mitochondrial membrane potential, and by evaluating its apoptotic induction potential. The cell viability of breast cancer cells was drastically decreased by vanillic acid, and its antiproliferative potential in MCF-7 cells could be due to its ability to induce excessive generation of ROS and its apoptotic induction potential. The present investigation thus explores antioxidant and antiproliferative properties of vanillic acid under in vitro conditions.

Lysine metabolism is a novel metabolic tumor suppressor pathway in breast cancer.

The International Agency for Research on Cancer determined that obesity is the primary preventable cause of breast cancer. The nuclear receptor peroxisome proliferator activated receptor γ (PPARγ) binds inflammatory mediators in obesity and its expression is reduced in human breast cancer. We created a new model to better understand how the obese microenvironment alters nuclear receptor function in breast cancer. The obesity related cancer phenotype was PPARγ dependent; deletion of PPARγ in mammary epithelium which is a tumor suppressor in lean mice unexpectedly increased tumor latency, reduced the luminal progenitor (LP) tumor cell fraction, and increased autophagic and senescent cells. Loss of PPARγ expression in mammary epithelium of obese mice increased expression of 2-aminoadipate semialdehyde synthase (AASS) which regulates lysine catabolism to acetoacetate. PPARγ-associated co-repressors and activators regulated AASS expression via a canonical response element. AASS expression was significantly reduced in human breast cancer, and AASS overexpression or acetoacetate treatment inhibited proliferation and induced autophagy and senescence in human breast cancer cell lines. Genetic or pharmacologic HDAC inhibition promoted autophagy and senescence in mammary tumor cells in vitro and in vivo. We concluded that lysine metabolism is a novel metabolic tumor suppressor pathway in breast cancer.