Effector CD4+ T cell subsets in Takayasu arteritis - Differences between the peripheral blood and the aorta.

Takayasu arteritis (TAK) is a granulomatous vasculitis that affects large arteries. T cells are important in TAK pathophysiology as these cells orchestrate granulomatous infiltration in arteries. This study aims to evaluate effector CD4+ T cells in the peripheral blood and the aortic wall of TAK patients and to analyze associations with disease activity and therapy. We performed a longitudinal study including 30 TAK patients and 30 controls. CD3+ T cells, CD3+CD4- T cells, CD4+ T cells, and Th1, Th2, and Th17 cells were evaluated in peripheral blood by flow cytometry, and the expression of CD4, CD8, Tbet, GATA-3, and RORγT was analyzed in the aorta of 6 patients by immunohistochemistry. TAK patients presented lower CD3+ T cells and CD4+ T cells (p=0.031 and p=0.039, respectively) than controls. Patients with active disease and those in remission had higher proportions of Th17 cells than controls (p=0.016 and p=0.004, respectively). Therapy for TAK did not result in significant differences concerning CD4+ effector T cell subpopulations. Disease duration correlated with the number and percentage of Th2 cells (rho=-0.610 and rho=-0.463, respectively) and with Th17 cells (rho=-0.365 and rho=-0.568). In the aorta, the expression of CD8 was higher than CD4, whereas GATA-3, Tbet and RORγT were expressed in this order of frequency. In conclusion, TAK patients present an increased Th17 response in the peripheral blood regardless of disease activity, whereas in the aortic tissue CD8 cells and the Th2 response were predominant.

Recent Trends in Curcumin-Containing Inorganic-Based Nanoparticles Intended for In Vivo Cancer Therapy.

Curcumin is a natural compound that has been widely investigated thanks to its various biological properties, including antiproliferative. This molecule acts on different cancers such as lung, breast, pancreatic, colorectal, etc. However, the bioactive actions of curcumin have limitations when its physicochemical properties compromise its pharmacological potential. As a therapeutic strategy against cancer, curcumin has been associated with inorganic nanoparticles. These nanocarriers are capable of delivering curcumin and offering physicochemical properties that synergistically enhance anticancer properties. This review highlights the different types of curcumin-based inorganic nanoparticles and discusses their physicochemical properties and in vivo anticancer activity in different models of cancer.

Biomarkers of oxidative stress and inflammation in subjects with COVID-19: Characterization and prognosis of the disease.

Coronavirus disease (COVID-19) is an acute respiratory disease caused by the new coronavirus (SARS-CoV-2) that has spread throughout the world causing millions of deaths. COVID-19 promotes excessive release of pro-inflammatory cytokines leading to acute lung injury and death. Reactive oxygen species (ROS) and oxidative stress (OS) may also play a role in the pathophysiology of COVID-19. The present study investigated levels of inflammatory cytokines (IL-1ß, IL-6, IL-8, IL-10, IL-12) and OS biomarkers (MPO, SOD, CAT, GST enzymes and contents of GSH, TBARS and PC) in patients with SARS-CoV-2 infection, which were correlated with disease severity. Patients with SARS significantly increased IL-1ß levels, while IL-6 levels were elevated in both groups of SARS-CoV-2 positive patients. The most severe patients showed increased levels of IL-8 and IL-10, while subjects without SARS showed lower values. MPO activity were higher in both groups of SARS-CoV-2 positive patients, while SOD and CAT activity were decreased in both groups. Compared to controls, GGT was elevated only in the SARS patient group, while GST values were increased in the group of positive patients in SARS-CoV-2 without SARS and were decreased in patients with SARS. GSH and UA contents decreased in SARS-CoV-2 positive subjects, whereas TBARS and PC contents increased in both groups of SARS-CoV-2 positive patients, particularly in the SARS patient group. In addition, several important correlations were found between cytokines and the different OS parameters suggesting some inter-relationship in the complex antioxidant system of the patients. In general, patients with SARS-CoV-2 infection showed higher levels of OS biomarkers, and also elevated contents of IL-6 and IL-10, probably worsening the damage caused by SARS-CoV-2 infection. This damage may contribute to the severity of the disease and its complications, as well as a prognosis for SARS-CoV-2 patients.

Protein restriction impairs the response activation/responsivity of MAPK signaling pathway of hematopoietic stem cells.

Protein restriction (PR) leads to bone marrow hypoplasia with changes in stromal cellularity components of the extracellular matrix in hematopoietic stem cells (HSCs). However, the underlying signaling mechanisms are poorly understood. We hypothesize that PR impairs the HSC mitogen-activated protein kinase (MAPK) signaling pathway response activation. Our aim is to evaluate the activation of MAPK and interleukin-3 (IL-3) proteins in HSC to explain PR-induced bone marrow hypoplasia, which causes altered proliferation and differentiation. C57BL/6 male mice were subjected to a low-protein diet (2% protein) or normoproteic (12% protein). PKC, PLCγ2, CaMKII, AKT, STAT3/5, ERK1/2, JNK, and p38d phosphorylation were evaluated by flow cytometry, and GATA1/2, PU.1, C/EBPα, NF-E2, and Ikz-3 genes (mRNAs) assessed by quantitative real-time-polymerase chain reaction. Pathway proteins, such as PLCγ2, JAK2, STAT3/5, PKC, and RAS do not respond to the IL-3 stimulus in PR, leading to lower activation of ERK1/2 and Ca2+ signaling pathways, consequently lowering the production of hematopoietic transcription factors. Colony forming units granulocyte-macrophage and colony forming units macrophage formation are impaired in PR even after being stimulated with IL-3. Long-term hematopoietic stem cells, short-term hematopoietic stem cells, granulocyte myeloid progenitor, and megakaryocyte-erythroid progenitor cells were significantly reduced in PR animals. This study shows for the first time that activation of MAPK pathway key proteins in HSCs is impaired in cases of PR. Several pathway proteins, such as PLCγ2, JAK2, STAT3, PKC, and RAS do not respond to IL-3 stimulation, leading to lower activation of extracellular signal-regulated protein kinase 1/2 and consequently lower production of hematopoietic transcription factors GATA1/2, PU.1, C/EBPa, NF-E2, and Ikz3. These changes result in a reduction in colony-forming units, proliferation, and differentiation, leading to hypocellularity.

Monocyte subsets and monocyte-related chemokines in Takayasu arteritis.

The pathogenesis of Takayasu arteritis (TAK) is poorly understood and no previous studies have analyzed monocytes in TAK. This study evaluated monocyte subsets and monocyte-related chemokines in the peripheral blood of TAK patients and healthy controls (HC). Monocyte subsets were identified as classical (CD14+CD16-), intermediate (CD14+CD16dim), and non-classical (CD14dimCD16high) in the peripheral blood. The chemokines CCL (C-C chemokine ligand)2, CCL3, CCL4, CCL5, CCL7, CXCL (C-X-C motif ligand)10, and CX3CL (C-X3-C motif ligand)1 were measured in the sera. Thirty-two TAK patients and 30 HC were evaluated. Intermediate monocytes were higher in TAK than HC [25.0 cells ×106/L (16.7-52.0) vs. 17.2 cells ×106/L (9.2-25.3); p = 0.014]. Active disease was associated with monocytosis (p = 0.004), increased classical (p = 0.003), and intermediate (p < 0.001) subsets than HC. Prednisone reduced the percentage of non-classical monocytes (p = 0.011). TAK patients had lower CCL3 (p = 0.033) and CCL4 (p = 0.023) levels than HC, whereas CCL22 levels were higher in active TAK compared to the remission state (p = 0.008). Glucocorticoids were associated with lower CXCL10 levels (p = 0.012). In TAK, CCL4 correlated with total (Rho = 0.489; p = 0.005), classical and intermediate monocytes (Rho = 0.448; p = 0.010 and Rho = 0.412; p = 0.019). In conclusion, TAK is associated with altered counts of monocyte subsets in the peripheral blood compared to HC and CCL22 is the chemokine with the strongest association with active disease in TAK.

Selenylated Imidazo[1,2-a]pyridine Induces Cell Senescence and Oxidative Stress in Chronic Myeloid Leukemia Cells.

Imidazo[1,2-a]pyridines (IPs) have been studied regarding drug development. The objective of this work was to evaluate the antileukemic capacity of IP derivatives by screening their ability as a pro-oxidant. IP derivatives were synthesized and oral bioavailability and toxicity were analyzed in silico. Redox screening was performed on human Kasumi, KG-1, K562, and Jurkat leukemia cells. The IP derivative and the most responsive leukemic cell were selected for cytotoxicity, cell proliferation, cell senescence, and oxidative stress assays. The predictive toxicity analysis showed a possible effect on the reproductive system, but without mutagenic, carcinogenic, or irritability effects. MRK-107 against K562 cells was the compound that showed the best redox profile. MRK-107 did not induce cell death in K562 and monocyte cells. However, this compound was able to decrease cell proliferation and increase cell senescence after 48 and 72 h. Furthermore, MRK-107 induced oxidative stress in K562 cells after 72 h, increasing lipid peroxidation and decreasing reduced glutathione (GSH) contents. This study demonstrated that MRK-107-induced senescence with the involvement of oxidative stress is a possible mechanism of action, addressing this compound as a potential antitumor drug against chronic myeloid leukemia.

In Vitro Study of Cytotoxic Mechanisms of Alkylphospholipids and Alkyltriazoles in Acute Lymphoblastic Leukemia Models.

This study investigates the efficacy of miltefosine, alkylphospholipid, and alkyltriazolederivative compounds against leukemia lineages. The cytotoxic effects and cellular and molecular mechanisms of the compounds were investigated. The inhibitory potential and mechanism of inhibition of cathepsins B and L, molecular docking simulation, molecular dynamics and binding free energy evaluation were performed to determine the interaction of cathepsins and compounds. Among the 21 compounds tested, C9 and C21 mainly showed cytotoxic effects in Jurkat and CCRF-CEM cells, two human acute lymphoblastic leukemia (ALL) lineages. Activation of induced cell death by C9 and C21 with apoptotic and necrosis-like characteristics was observed, including an increase in annexin-V+propidium iodide-, annexin-V+propidium iodide+, cleaved caspase 3 and PARP, cytochrome c release, and nuclear alterations. Bax inhibitor, Z-VAD-FMK, pepstatin, and necrostatin partially reduced cell death, suggesting that involvement of the caspase-dependent and -independent mechanisms is related to cell type. Compounds C9 and C21 inhibited cathepsin L by a noncompetitive mechanism, and cathepsin B by a competitive and noncompetitive mechanism, respectively. Complexes cathepsin-C9 and cathepsin-C21 exhibited significant hydrophobic interactions, water bridges, and hydrogen bonds. In conclusion, alkyltriazoles present cytotoxic activity against acute lymphoblastic lineages and represent a promising scaffold for the development of molecules for this application.

First generation of multifunctional peptides derived from latarcin-3a from Lachesana tarabaevi spider toxin.

The need for discovering new compounds that can act selectively on pathogens is becoming increasingly evident, given the number of deaths worldwide due to bacterial infections or tumor cells. New multifunctional biotechnological tools are being sought, including compounds present in spider venoms, which have high biotechnological potential. The present work aims to perform the rational design and functional evaluation of synthetic peptides derived from Lachesana tarabaevi spider toxin, known as latarcin-3a. The antimicrobial activity was tested against Gram-positive and -negative bacteria, with minimum inhibitory concentrations (MIC) between 4 and 128 µg.ml-1. Anti-biofilm tests were then performed to obtain MICs, where the peptides demonstrated activity from 4 to 128 µg.ml-1. In vitro cell cytotoxicity assays were carried out from tumor cell lines, lineages C1498, Kasumi-1, K-562, Jurkat, MOLT4, and Raji. Erythrocyte integrity was evaluated in the presence of synthetic peptides analog, which did not promote hemolysis at 128 µg.ml-1. The peptide that showed the best antibacterial activity was Lt-MAP3 and the best antitumor was Lt-MAP2. In conclusion, rational design of multifunctional antimicrobial peptides may be promising alternative tools in the treatment of emerging diseases such as bacterial infections and tumor cells.

Effect of Ylang-Ylang (Cananga odorata Hook. F. & Thomson) Essential Oil on Acute Inflammatory Response In Vitro and In Vivo.

The aim of this study is to evaluate the phytochemical profile, oral acute toxicity, and the effect of ylang-ylang (Cananga odorata Hook. F. & Thomson) essential oil (YEO) on acute inflammation. YEO was analyzed by gas chromatography/mass spectrometry. For in vitro tests, YEO was assessed using cytotoxicity, neutrophil chemotaxis induced by N-formyl methionyl leucyl phenylalanine (fMLP), and phagocytic activity tests. YEO was orally administered in zymosan-induced peritonitis, carrageenan-induced leukocyte rolling, and adhesion events in the in situ microcirculation model and in carrageenan-induced paw edema models. YEO (2000 mg/kg) was also tested using an acute toxicity test in Swiss mice. YEO showed a predominance of benzyl acetate, linalool, benzyl benzoate, and methyl benzoate. YEO did not present in vitro cytotoxicity. YEO reduced the in vitro neutrophil chemotaxis induced by fMLP and reduced the phagocytic activity. The oral treatment with YEO reduced the leukocyte recruitment and nitric oxide production in the zymosan-induced peritonitis model, reduced rolling and adherent leukocyte number induced by carrageenan in the in situ microcirculation model, and reduced carrageenan-induced edema and mechanical hyperalgesia. YEO did not present signs of toxicity in the acute toxicity test. In conclusion, YEO affected the leukocyte activation, and presented antiedematogenic, anti-hyperalgesic, and anti-inflammatory properties.

Direct ionizing radiation and bystander effect in mouse mesenchymal stem cells.

Purpose: This study aimed to evaluate the radiation-induced direct and bystander (BYS) responses of mesenchymal stem cells (MSCs) and to characterize these cells radiobiologically.Methods and materials: MSCs were irradiated (IR) and parameters related to DNA damage and cellular signaling were verified in a dose range from 0.5 to 15 Gy; also a transwell insert co-culture system was used to study medium-mediated BYS effects.Results: The main effects on directly IR cells were seen at doses higher than 6 Gy: induction of cell death, cell cycle arrest, upregulation of p21, and alteration of redox status. Irrespective of a specific dose, induction of micronuclei formation, H2AX phosphorylation, and decreased Akt expression also occurred. Thus, mTOR expression, cell senescence, nitric oxide generation, and calcium levels, in general were not significantly modulated by radiation. Data from the linear-quadratic model showed a high alpha/beta ratio, which is consistent with a more exponential survival curve. BYS effects from the unirradiated MSCs placed into companion wells with the directly IR cells, were not observed.Conclusions: The results can be interpreted as a positive outcome, meaning that the radiation damage is restricted to the directed IR MSCs not leading to off-target cell responses.

Chemodiversity and Anti-Leukemia Effect of Metabolites from Penicillium setosum CMLD 18.

Penicillium setosum represents a Penicillium species recently described, with little up-to-date information about its metabolic and biological potential. Due to this scenario, we performed chemical and biological studies of P. setosum CMLD18, a strain isolated from Swinglea glutinosa (Rutaceae). HRMS-MS guided dereplication strategies and anti-leukemia assays conducted the isolation and characterization of six compounds after several chromatographic procedures: 2-chloroemodic acid (2), 2-chloro-1,3,8-trihydroxy-6- (hydroxymethyl)-anthraquinone (7), 7-chloroemodin (8), bisdethiobis(methylthio)acetylaranotine (9), fellutanine C (10), and 4-methyl-5,6-diihydro-2H-pyran-2-one (15). From the assayed metabolites, (10) induced cellular death against Kasumi-1, a human leukemia cell line, as well as good selectivity for it, displaying promising cytotoxic activity. Here, the correct NMR signal assignments for (9) are also described. Therefore, this work highlights more detailed knowledge about the P. setosum chemical profile as well as its biological potential, offering prospects for obtaining natural products with anti-leukemia capabilities.

Distribution of monocytes subpopulations in the peripheral blood from patients with Behçet's disease - Impact of disease status and colchicine use.

The innate immune response has a predominant role in Behçet's disease (BD) pathogenesis, but few studies have assessed monocytes in BD. This study aims to evaluate the profile of monocytes subsets in the peripheral blood of BD patients and healthy controls (HC). Monocytes subsets were identified as classical (CD14+CD16-), intermediate (CD14+CD16dim), and non-classical (CD14dimCD16high) subsets. Patients with BD presented a lower number of total monocytes (p = 0.020) and a lower number (p < 0.0001) of circulating classical monocytes than HC. In contrast, the number of intermediate monocytes was higher in BD patients than HC (p < 0.0001). In BD patients, no associations were observed with the severity of clinical manifestations or therapy. Colchicine was associated with a higher number of non-classical monocytes (p = 0.035). In conclusion, BD patients present an altered distribution of monocytes subsets with a reduction of classical and an increase of intermediate subsets.

α-amyrin-loaded nanocapsules produce selective cytotoxic activity in leukemic cells.

INTRODUCTION: Amyrins are triterpenes that have attractive pharmacological potential; however, their low water solubility and erratic stomach absorption hinders their use as a drug. The aim of this paper was to develop a novel α-amyrin-loaded nanocapsule for intestinal delivery and evaluate, preliminarily, its cytotoxic ability against leukemic cells. MATERIAL AND METHODS: Five nanocapsule formulations were designed by the solvent displacement-evaporation method. Poly-ε-caprolactone, Eudragit® E100, and Kollicoat® Mae 100 P were used as film-former materials. Particle size, polydispersity index (PdI), zeta potential, and the pH of all formulations were measured. The cytotoxic potential of the nanocapsules was evaluated in vitro using different leukemic lineages RESULTS: Nanocapsules coated with Kollicoat® Mae 100 P presented the smallest particle size (130 nm), the lowest zeta-potential (-38 mV), and the narrowest size distribution (PdI = 0.100). The entrapment efficiency was 65.47%, while the loading capacity was 2.40%. Nanocapsules release 100% of α-amyrin in 40 min (pH 7.4), by using a possible mechanism of swelling-diffusion. The formulation showed excellent on-shelf physicochemical stability during one year. Additionally, nanocapsules produced a selective cytotoxic effect on a human leukemia lineage Kasumi-1, an acute myeloid leukemia cell line, and produced cell death by apoptosis CONCLUSION: α-amyrin-loaded nanocapsules appear to be a promising nanoformulation that could be used against leukemia.

A styrylpyrone dimer isolated from Aniba heringeri causes apoptosis in MDA-MB-231 triple-negative breast cancer cells.

The styrylpyrone dehydrogoniothalamin (1) and two of its dimers (2 and 3) were isolated from the leaves of Aniba heringeri (Lauraceae). Compound 3 is new, while 1 and 2 are being reported for the first time in this species. Structures were determined by 1D- and 2D-NMR spectroscopy, mass spectrometry, and optical rotation data. Cytotoxic effects and selectivity indices were evaluated in five neoplastic cell lines-PC-3 (prostate), 786-0 (renal), HT-29 (colon), MCF-7, and MDA-MB-231 (breast)-and a non-neoplastic cell line, (NIH/3T3, murine fibroblast). Compound 1 inhibited cell growth by 50% (GI50) at concentrations in the 90.4-175.7 µM range, while 2 proved active against MCF-7 and MDA-MB-231 breast cells (GI50 = 12.24, and 34.22 µM, respectively). Compound 3 showed strong cytotoxicity (GI50 = 4.4 µM) against MDA-MB-231 (an established basal triple-negative breast carcinoma (TNBC) cell line), with a high selective index of 35. This compound was subsequently evaluated for apoptosis induction in MDA-MB-231 cells, using GI50 and 50% lethal concentrations (LC50). Flow cytometry analysis showed that at LC50 compound 3 induced cell death with phosphatidylserine externalization and caspase-3 activation. Apoptotic genes were measured by RT-qPCR, revealing an upregulation of BAX, with an increase in expression of the BAX/BCL2 ratio in treated cells. Fluorescence microscopy disclosed morphological changes related to apoptosis. Overall, these findings showed compound 3 to be a promising prototype against TNBC cells that tend to respond poorly to conventional therapies.

Gemfibrozil Induces Anemia, Leukopenia and Reduces Hematopoietic Stem Cells via PPAR-α in Mice.

Hypercholesterolemia, also called high cholesterol, is a form of hyperlipidemia, which may be a consequence of diet, obesity or diabetes. In addition, increased levels of low-density lipoprotein (LDL) and reduced levels of high-density lipoprotein (HDL) cholesterol are associated with a higher risk of atherosclerosis and coronary heart disease. Thus, controlling cholesterol levels is commonly necessary, and fibrates have been used as lipid-lowering drugs. Gemfibrozil is a fibrate that acts via peroxisome proliferator-activated receptor alpha to promote changes in lipid metabolism and decrease serum triglyceride levels. However, anemia and leukopenia are known side effects of gemfibrozil. Considering that gemfibrozil may lead to anemia and that gemfibrozil acts via peroxisome proliferator-activated receptor alpha, we treated wild-type and peroxisome proliferator-activated receptor alpha-knockout mice with gemfibrozil for four consecutive days. Gemfibrozil treatment led to anemia seven days after the first administration of the drug; we found reduced levels of hemoglobin, as well as red blood cells, white blood cells and a reduced percentage of hematocrits. PPAR-alpha-knockout mice were capable of reversing all of those reduced parameters induced by gemfibrozil treatment. Erythropoietin levels were increased in the serum of gemfibrozil-treated animals, and we also observed an increased expression of hypoxia-inducible factor-2 alpha (HIF-2α) and erythropoietin in renal tissue, while PPAR-alpha knockout mice treated with gemfibrozil did not present increased levels of serum erythropoietin or tissue HIF-2α and erythropoietin mRNA levels in the kidneys. We analyzed bone marrow and found that gemfibrozil reduced erythrocytes and hematopoietic stem cells in wild-type mice but not in PPAR-alpha-knockout mice, while increased colony-forming units were observed only in wild-type mice treated with gemfibrozil. Here, we show for the first time that gemfibrozil treatment leads to anemia and leukopenia via peroxisome proliferator-activated receptor alpha in mice.

Antimicrobial and antileukemic effects: in vitro activity of Calyptranthes grandifolia aqueous leaf extract.

Natural products are still a promising source of bioactive molecules. Food and Drug Administration data showed that approximately 49% of the approved molecules originate naturally or chemically-resemble these substances, of which more than 70% are being used in anticancer therapy. It is noteworthy that at present there are no scientific studies to prove the effectiveness and safety of a number of plants used in folk medicine such as in the case of Calyptranthes grandifolia O. Berg (Myrtaceae) originally from South America. The aim of the present study was to determine the biological potential and toxicological effects of the aqueous leaf extract of C. grandifolia. The main detected phytoconstituents were condensed tannins and flavonoids and a high quantity of polyphenols. Regarding the antimicrobial potential, the extract exerted inhibitory activity against Pseudomonas aeruginosa. The results also revealed the extract induced DNA damage in a concentration-dependent manner in RAW 264.7 cells. In addition, C. grandifolia produced cytotoxicity in leukemia cell lines (HL60 and Kasumi-1) without affecting isolated human lymphocytes but significantly inhibited JAK3 and p38α enzyme activity. Taken together, these findings add important information on the biological and toxicological effects of C. grandifolia, indicating that aqueous extract may be a source of natural antimicrobial and antileukemic constituents.

Divergent effects of Wnt5b on IL-3- and GM-CSF-induced myeloid differentiation.

The multiple specialized cell types of the hematopoietic system originate from differentiation of hematopoietic stem cells and progenitors (HSPC), which can generate both lymphoid and myeloid lineages. The myeloid lineage is preferentially maintained during ageing, but the mechanisms that contribute to this process are incompletely understood. Here, we studied the roles of Wnt5a and Wnt5b, ligands that have previously been linked to hematopoietic stem cell ageing and that are abundantly expressed by both hematopoietic progenitors and bone-marrow derived niche cells. Whereas Wnt5a had no major effects on primitive cell differentiation, Wnt5b had profound and divergent effects on cytokine-induced myeloid differentiation. Remarkably, while IL-3-mediated myeloid differentiation was largely repressed by Wnt5b, GM-CSF-induced myeloid differentiation was augmented. Furthermore, in the presence of IL-3, Wnt5b enhanced HSPC self-renewal, whereas in the presence of GM-CSF, Wnt5b accelerated differentiation, leading to progenitor cell exhaustion. Our results highlight discrepancies between IL-3 and GM-CSF, and reveal novel effects of Wnt5b on the hematopoietic system.

AMPK activation induced by promethazine increases NOXA expression and Beclin-1 phosphorylation and drives autophagy-associated apoptosis in chronic myeloid leukemia.

Relapse and drug resistance is still major challenges in the treatment of leukemia. Promethazine, an antihistaminic phenothiazine derivative, has been used to prevent chemotherapy-induced emesis, although there is no report about its antitumor potential. Thus, we evaluated the promethazine cytotoxicity against several leukemia cells and the underlying mechanisms were investigated. Promethazine exhibited potent and selective cytotoxicity against all leukemia cell types in vitro at clinically relevant concentrations. Philadelphia positive chronic myeloid leukemia (CML) K562 cells were the most sensitive cell line. The cytotoxicity of promethazine in these cells was triggered by the activation of AMPK and inhibition of PI3K/AKT/mTOR pathway. The subsequent downstream effects were NOXA increase, MCL-1 decrease, and Beclin-1 activation, resulting in autophagy-associated apoptosis. These data highlight targeting autophagy may represent an interesting strategy in CML therapy, and also the antitumor potential of promethazine by acting in AMPK and PI3K/AKT/mTOR signaling pathways. Since this drug is currently used with relative low side effects, its repurposing may represent a new therapeutic opportunity for leukemia treatment.

Ethanolic Extract of Senna velutina Roots: Chemical Composition, In Vitro and In Vivo Antitumor Effects, and B16F10-Nex2 Melanoma Cell Death Mechanisms.

Cutaneous melanoma is among the most aggressive types of cancer, and its rate of occurrence increases every year. Current pharmacological treatments for melanoma are not completely effective, requiring the identification of new drugs. As an alternative, plant-derived natural compounds are described as promising sources of new anticancer drugs. In this context, the objectives of this study were to identify the chemical composition of the ethanolic extract of Senna velutina roots (ESVR), to assess its in vitro and in vivo antitumor effects on melanoma cells, and to characterize its mechanisms of action. For these purposes, the chemical constituents were identified by liquid chromatography coupled to high-resolution mass spectrometry. The in vitro activity of the extract was assessed in the B16F10-Nex2 melanoma cell line using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and based on the apoptotic cell count; DNA fragmentation; necrostatin-1 inhibition; intracellular calcium, pan-caspase, and caspase-3 activation; reactive oxygen species (ROS) levels; and cell cycle arrest. The in vivo activity of the extract was assessed in models of tumor volume progression and pulmonary nodule formation in C57Bl/6 mice. The chemical composition results showed that ESVR contains flavonoid derivatives of the catechin, anthraquinone, and piceatannol groups. The extract reduced B16F10-Nex2 cell viability and promoted apoptotic cell death as well as caspase-3 activation, with increased intracellular calcium and ROS levels as well as cell cycle arrest at the sub-G0/G1 phase. In vivo, the tumor volume progression and pulmonary metastasis of ESVR-treated mice decreased over 50%. Combined, these results show that ESVR had in vitro and in vivo antitumor effects, predominantly by apoptosis, thus demonstrating its potential as a therapeutic agent in the treatment of melanoma and other types of cancer.

Changes in human intervertebral disc biochemical composition and bony end plates between middle and old age.

OBJECTIVE: This study evaluates molecular, nutritional and biochemical alterations in human intervertebral discs between middle and old age. METHODS: Twenty-eight human lumbar intervertebral discs from donors were evaluated and separated into two groups: Middle-aged (35-50 years old, relatively non-degenerate discs of Pfirrmann grades 1-3, n = 15) and Old-aged (≥80 years old, all degenerate Pfirrmann grade 4 or 5, n = 13). Parameters which might be expected to to be related to nutrient supply and so the health of disc cells (eg the porosity of the vertebral endplate, cell viability and cell density) and to disc extracellular composition (ie quantification of glycosaminoglycan disaccharides and hyaluronic acid molecular weight) and collagen organization, were analyzed. Three regions of the intervertebral disc (anterior annulus fibrosus, nucleus pulposus, and posterior annulus fibrosus) were examined. RESULTS: The old-aged group showed a decrease in content of sulphated and non-sulphated glycosaminoglycans relative to middle-aged and there were also alterations in the proportion of GAG disaccharides and a decrease of collagen fiber size. Hyaluronic acid molecular weight was around 200 kDa in all regions and ages studied. The anterior annulus differed from the posterior annulus particularly in relation to cell density and GAG content. Additionally, there were changes in the bony endplate, with fewer openings observed in the caudal than cranial endplates of all discs in both groups. CONCLUSIONS: Results show the cranial vertebral endplate is the main vascular source for the intervertebral discs. Hylauronic acid molecular weight is the same through the intervertebral disc after age of 50 years.