UNASSIGNED: To investigate serum cholesterol efflux capacity (the ability of the serum to accept cholesterol) and factors that regulate it using nuclear magnetic resonance-quantified measures of lipoprotein particle composition and size and apolipoproteins metrics in patients with age-related macular degeneration (AMD).
UNASSIGNED: Case-control study.
UNASSIGNED: Four hundred two serum samples from 80 patients with early AMD (eAMD), and 212 patients with neovascular AMD (nAMD), including 80 with typical nAMD (tAMD) and 132 with polypoidal choroidal vasculopathy (PCV), and 110 age- and gender matched control participants.
UNASSIGNED: Serum from participants showed cholesterol efflux capacity measured using in vitro cell assays and lipoprotein subfractions measured using nuclear magnetic resonance (Nightingale, Ltd). Associations between cholesterol efflux capacity (measured in percentage) and lipid subfractions were investigated in the patients and control participants.
UNASSIGNED: Cholesterol efflux capacity and lipid subfractions in control, eAMD, and nAMD. Associations between HDL subfractions and cholesterol efflux capacity.
UNASSIGNED: Cholesterol efflux capacity was higher in patients with eAMD (68.0 ± 11.3% [mean ± standard deviation]) and nAMD (75.9 ± 27.7%) than in the control participants (56.9 ± 16.7%) after adjusting for age, gender, and use of lipid-lowering drug (P < 0.0001). Nuclear magnetic resonance lipidomics demonstrated that the mean diameter of HDL was larger both in eAMD (9.96 ± 0.27 mm [mean ± standard deviation]) and PCV (9.97 ± 0.23 mm) compared with that of the control participants (9.84 ± 0.24 mm; P = 0.0001 for both). Among the 28 HDL subfractions, most of the small, medium, and large HDLs, but none of the 7 extra large HDLs fractions, were associated moderately with cholesterol efflux capacity in eAMD and PCV (R = 0.149-0.277).
UNASSIGNED: Serum cholesterol efflux capacity was increased in eAMD and PCV, but not tAMD, possibly reflecting differential underlying pathophysiologic features of lipid dysregulation in tAMD and PCV. Further studies should be directed toward investigating the diverse biological activities of HDL in AMD, including macular pigment transport, regulation of inflammation, and local cholesterol transport system.

Neomenthol, a cyclic monoterpenoid, is a stereoisomer of menthol present in the essential oil of Mentha spp. It is used in food as a flavoring agent, in cosmetics and medicines because of its cooling effects. However, neomenthol has not been much explored for its anticancer potential. Additionally, targeting hyaluronidase, Cathepsin-D, and ODC by phytochemicals is amongst the efficient approach for cancer prevention and/or treatment.
To investigate the molecular and cell target-based antiproliferative potential of neomenthol on human cancer (A431, PC-3, K562, A549, FaDu, MDA-MB-231, COLO-205, MCF-7, and WRL-68) and normal (HEK-293) cell lines.
The potency of neomenthol was evaluated on human cancer and normal cell line using SRB, NRU and MTT assays. The molecular target based study of neomenthol was carried out in cell-free and cell-based test systems. Further, the potency of neomenthol was confirmed by quantitative real-time PCR analysis and molecular docking studies. The in vivo anticancer potential of neomenthol was performed on mice EAC model and the toxicity examination was accomplished through in silico, ex vivo and in vivo approaches.
Neomenthol exhibits a promising activity (IC50 17.3 ± 6.49 μM) against human epidermoid carcinoma (A431) cells by arresting the G2/M phase and increasing the number of sub-diploid cells. It significantly inhibits hyaluronidase activity (IC50 12.81 ± 0.01 μM) and affects the tubulin polymerization. The expression analysis and molecular docking studies support the in vitro molecular and cell target based results. Neomenthol prevents EAC tumor formation by 58.84% and inhibits hyaluronidase activity up to 10% at 75 mg/kg bw, i.p. dose. The oral dose of 1000 mg/kg bw was found safe in acute oral toxicity studies.
Neomenthol delayed the growth of skin carcinoma cells by inhibiting the tubulin polymerization and hyaluronidase activity, which are responsible for tumor growth, metastasis, and angiogenesis.

A Newcastle disease virus (NDV) oncolysate has been established as a unique and effective immune-stimulatory root for tumor treatment. Thus, the aim of the current study was to investigate the effects of intratumoral administration of NDV oncolysate on immune response and tumor regression of C57BL/6 mouse model of human papillomavirus (HPV) related transplanted with TC-1 syngeneic cancer cells. To further investigate the mechanism underlying the antitumor response, cytolytic and lymphocyte proliferation responses in splenocytes were measured using lactate dehydrogenase (LDH) release and MTT assays, respectively. In this regard, levels of IL-10, IFN-γ, and IL-4 were measured using ELISA after re-stimulation. The immune responses efficacy was evaluated by in vivo tumor regression assay. The results showed that immunization with the different titers of NDV lysate significantly reduced tumor volume in comparison with a combination of virus lysate and tumor cell lysate. Also, virus lysate could significantly enhance cytotoxic T lymphocyte production and lymphocyte proliferation rates versus tumor cell lysate. Also, our major findings are that the peritumorally injection of NDV oncolysate effectively induces antitumor immune responses through increased levels of IL-4, IFN-γ, and reduction of IL-10. These results indicate that this treatment is a specific, active immune mechanism stimulator, and may prove to be a useful therapeutic for a treatment against cervical cancers and merits further investigation.

Rhabdomyosarcoma (RMS) is the most common pediatric soft-tissue malignant tumor. Treatment of RMS usually includes primary tumor resection along with systemic chemotherapy. Two-dimensional (2D) cell culture systems and animal models have been extensively used for investigating the potential efficacy of new RMS treatments. However, RMS cells behave differently in 2D culture than in vivo, which has recently inspired the adoption of three-dimensional (3D) culture environments. In the current paper, we will describe the detailed methodology we have developed for fabricating a 3D engineered model to study alveolar RMS (ARMS) in vitro. This model consists of a thermally cross-linked collagen disk laden with RMS cells that mimics the structural and bio-chemical aspects of the tumor extracellular matrix (ECM). This process is highly reproducible and produces a 3D engineered model that can be used to analyze the cytotoxicity and autophagy induction of drugs on ARMS cells. The most improtant bullet points are as following:•We fabricated 3D model of ARMS.•The current ARMS 3D model can be used for screening of chemotherapy drugs.•We developed methods to detect apoptosis and autophagy in ARMS 3D model to detect the mechansims of chemotherapy agents.

Trypanosoma cruzi is the agent of Chagas disease, an infection that affects around 8 million people worldwide. The search for new anti-T. cruzi drugs are relevant, mainly because the treatment of this disease is limited to two drugs. The objective of this study was to investigate the trypanocidal and cytotoxic activity and elucidate the chemical profile of extracts from the roots of the Lonchocarpus cultratus. Roots from L. cultratus were submitted to successive extractions with hexane, dichloromethane, and methanol, resulting in LCH, LCD, and LCM extracts, respectively. Characterization of extracts was done using 1H-RMN, 13C-RMN, CC and TLC. Treatment of T. cruzi forms (epimastigotes, trypomastigotes, and amastigotes) with crescent concentrations of LCH, LCD, and LCM was done for 72, 48, and 48 h, respectively. After this, the percentage of inhibition and IC50/LC50 were calculated. Benznidazole was used as a positive control. Murine macrophages were treated with different concentrations of both extracts for 48 h, and after, the cellular viability was determined by the MTT method and CC50 was calculated. The chalcones derricin and lonchocarpine were identified in the hexane extract, and for the first time in the genus Lonchocarpus, the presence of a dihydrolonchocarpine derivative was observed. Other chalcones such as isocordoin and erioschalcone B were detected in the dichloromethane extract. The dichloromethane extract showed higher activity against all tested forms of T. cruzi than the other two extracts, with IC50 values of 10.98, 2.42, and 0.83 µg/mL, respectively; these values are very close to those of benznidazole. Although the dichloromethane extract presented a cytotoxic effect against mammalian cells, it showed selectivity against amastigotes. The methanolic extract showed the lowest anti-T. cruzi activity but was non-toxic to peritoneal murine macrophages. Thus, the genus Lonchocarpus had demonstrated in the past action against epimastigotes forms of T. cruzi but is the first time that the activity against infective forms is showed, which leading to further studies with in vivo tests.

The genetic alterations associated with cell transformation are in large measure expressed in the metabolic phenotype as cancer cells proliferate and change their local environment, and prepare for metastasis. Qualitatively, the fundamental biochemistry of cancer cells is generally the same as in the untransformed cells, but the cancer cells produce a local environment, the TME, that is hostile to the stromal cells, and compete for nutrients. In order to proliferate, cells need sufficient nutrients, either those that cannot be made by the cells themselves, or must be made from simpler precursors. However, in solid tumors, the nutrient supply is often limiting given the potential for rapid proliferation, and the poor quality of the vasculature. Thus, cancer cells may employ a variety of strategies to obtain nutrients for survival, growth and metastasis. Although much has been learned using established cell lines in standard culture conditions, it is becoming clear from in vivo metabolic studies that this can also be misleading, and which nutrients are used for energy production versus building blocks for synthesis of macromolecules can vary greatly from tumor to tumor, and even within the same tumor. Here we review the operation of metabolic networks, and how recent understanding of nutrient supply in the TME and utilization are being revealed using stable isotope tracers in vivo as well as in vitro.

BACKGROUND: Formation of cell spheres is an important procedure in biomedical research. A large number of high-quality cell spheres of uniform size and shape are required for basic studies and therapeutic applications. Conventional approaches, including the hanging drop method and suspension culture, are used for cell sphere production. However, these methods are time consuming, cell spheres cannot be harvested easily, and it is difficult to control the size and geometry of cell spheres. To resolve these problems, a novel multiple-funnel cell culture insert was designed for size controlling, easy harvesting, and scale-up production of cell spheres.
METHODS: The culture substrate has 680 micro-funnels with a 1-mm width top, 0.89 mm depth, and 0.5 mm square bottom. Mouse embryonic stem cells were used to test the newly developed device. The seeded embryonic stem cells settled at the downward medium surface toward the bottom opening and aggregated as embryoid bodies (EBs). For cell sphere harvest, the bottom of the culture insert was put in contact with the medium surface in another culture dish, and the medium in the device flowed down with cell spheres by hydrostatic pressure.
RESULTS: Compact cell spheres with uniform size and shape were collected easily. The diameter of the spheres could be controlled by adjusting the seeding cell density. Spontaneous neural differentiation (nestin and Tju1) and retinoic acid-induced endodermal differentiation (Pdx-1 and insulin I) were improved in the EBs produced using the new insert compared to those in EBs produced by suspension culture.
CONCLUSIONS: This novel cell culture insert shall improve future studies of cell spheres and benefit clinical applications of cell therapy.

Certain food components possess immunomodulatory effects. The aim of this study was to elucidate the mechanism of the immunostimulatory activity of Brassica rapa L. We demonstrated an enhancement of natural killer (NK) activity and interferon (IFN)-γ production in mice that were orally administered an insoluble fraction of B. rapa L. The insoluble fraction of B. rapa L. significantly induced IFN-γ production in mouse spleen cells in an interleukin (IL)-12-dependent manner, and NK1.1+ cells were the main cells responsible for producing IFN-γ. Additionally, the results suggested that the active compounds in the insoluble fraction were recognized by Toll-like receptor (TLR) 2, TLR4, and C-type lectin receptors on dendritic cells, and they activated signaling cascades such as MAPK, NF-κB, and Syk. These findings suggest that B. rapa L. is a potentially promising immuno-improving material, and it might be useful for preventing immunological disorders such as infections and cancers by activating innate immunity.

OBJECTIVE: Mesalamine is a first-line drug for treatment of inflammatory bowel diseases (IBD). However, its mechanisms are not fully understood. CD4+ Foxp3+ regulatory T cells (Tregs) play a potential role in suppressing IBD. This study determined whether the anti-inflammatory activity of mesalamine is related to Treg induction in the colon.
METHODS: We examined the frequencies of Tregs in the colons of wild-type mice, mice deficient for aryl hydrocarbon receptor (AhR-/- mice), and bone marrow-chimeric mice lacking AhR in hematopoietic cells (BM-AhR-/- mice), following oral treatment with mesalamine. We also examined the effects of mesalamine on transforming growth factor (TGF)-β expression in the colon.
RESULTS: Treatment of wild-type mice with mesalamine increased the accumulation of Tregs in the colon and up-regulated the AhR target gene Cyp1A1, but this effect was not observed in AhR-/- or BM-AhR-/- mice. In addition, mesalamine promoted in vitro differentiation of naive T cells to Tregs, concomitant with AhR activation. Mice treated with mesalamine exhibited increased levels of the active form of TGF-β in the colon in an AhR-dependent manner and blockade of TGF-β signaling suppressed induction of Tregs by mesalamine in the colon. Furthermore, mice pretreated with mesalamine acquired resistance to dextran sodium sulfate-induced colitis.
CONCLUSIONS: We propose a novel anti-inflammatory mechanism of mesalamine for colitis: induction of Tregs in the colon via the AhR pathway, followed by TGF-β activation.

The anaplastic lymphoma kinase (ALK) fusion oncogene is observed in 3%-5% of non-small cell lung cancer (NSCLC). Crizotinib and ceritinib, a next-generation ALK tyrosine kinase inhibitor (TKI) active against crizotinib-refractory patients, are clinically available for the treatment of ALK-rearranged NSCLC patients, and multiple next-generation ALK-TKIs are currently under clinical evaluation. These ALK-TKIs exhibit robust clinical activity in ALK-rearranged NSCLC patients; however, the emergence of ALK-TKI resistance restricts the therapeutic effect. To date, various secondary mutations or bypass pathway activation-mediated resistance have been identified, but large parts of the resistance mechanism are yet to be identified. Here, we report the discovery of p-glycoprotein (P-gp/ABCB1) overexpression as a ceritinib resistance mechanism in ALK-rearranged NSCLC patients. P-gp exported ceritinib and its overexpression conferred ceritinib and crizotinib resistance, but not to PF-06463922 or alectinib, which are next-generation ALK inhibitors. Knockdown of ABCB1 or P-gp inhibitors sensitizes the patient-derived cancer cells to ceritinib, in vitro and in vivo. P-gp overexpression was identified in three out of 11 cases with in ALK-rearranged crizotinib or ceritinib resistant NSCLC patients. Our study suggests that alectinib, PF-06463922, or P-gp inhibitor with ceritinib could overcome the ceritinib or crizotinib resistance mediated by P-gp overexpression.