RESUMO
Metabolic reprogramming is a significant hallmark of cancer that promotes chemoresistance by allowing tumor tissues to adapt to changes in the tumor microenvironment caused by anticancer therapies. Hepatocellular carcinoma (HCC), one of the most common types of primary tumors, is associated with recurrent metabolic reprogramming that maximizes cancer cell growth and proliferation. Herein, we developed metformin (MET)-loaded hyaluronic acid (HA)-derived carbon dots (HA-CD-MET) by a simple and green method with no involvement of any additives. HA-CD-MET was utilized for specifically binding the CD44 receptor overexpressed in HCC and induced glutamine metabolic rewiring to inhibit HCC cell proliferation. Exposure to HA-CD-MET resulted in â¼6.5-fold better anticancer efficacy against CD44+ Hep3B cells in comparison to CD44-, HepG2, and noncancerous HEK293 cells at a very low dose of 80 µg/mL. Moreover, treatment of three-dimensional (3D) tumor spheroid model of HCC (Hep3B) with HA-CD-MET resulted in â¼4.9-fold reduction in tumor size. This improved anticancer efficacy of HA-CD-MET was attributed to the inhibition of glutaminase-1 (GLS-1), a mitochondrial enzyme that hydrolyzes glutamine into glutamate as confirmed from immunofluorescence and immunoblotting experiments. Furthermore, treatment with HA-CD-MET resulted in downregulation of glucose transporter-1 (GLUT-1) in Hep3B cells. Consequently, cancer cells were starved from essential nutrients, glutamine, and glucose, leading to the enhancement in intracellular ROS generation. This increase in intracellular ROS accumulation activated AMP-activated protein kinase (AMPK) and inhibited AKT phosphorylation, leading to cancer cell apoptosis. Thus, this study offers the targeting of metabolic reprogramming by HA-CD-MET that opens up a promising strategy for therapeutic intervention in hepatocarcinoma.
Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , Metformina , Humanos , Carcinoma Hepatocelular/patologia , Metformina/farmacologia , Metformina/uso terapêutico , Ácido Hialurônico , Glutamina , Neoplasias Hepáticas/patologia , Carbono , Espécies Reativas de Oxigênio/metabolismo , Células HEK293 , Linhagem Celular Tumoral , Receptores de Hialuronatos/metabolismo , Microambiente TumoralRESUMO
Fluorometric sensors have gained considerable attention in various fields, including environmental monitoring, biomedical research, and clinical diagnostics. This article delineates the fabrication of an orange emitting naphthalene diimide (NDI) derivative consisting of maleimide moiety (NDI-mal) for fluorometric sensing of thiols. Spherical shaped organic nanoparticles (â¼100-150 nm) were constructed by NDI-mal in dimethyl sulfoxide (DMSO)/dimethylformamide (DMF)-water through J-type aggregation. NDI-mal displayed self-assembly driven aggregation-induced emission (AIE) through excimer formation at λem= 588 nm at fw = 99 vol % DMSO/DMF-water. Naphthyl residue at both terminals of NDI-mal facilitates intramolecular charge transfer (ICT) from the donor naphthyl residue to the acceptor NDI core. The fluorescence intensity of NDI-mal fluorescent organic nanoparticles (FONPs) got quenched in the presence of thiols due to thiol-maleimide adduct formation (Michael addition). NDI-mal FONPs selectively probed thiol functionalized small molecules (4-aminothiophenol), biomolecules (glutathione (GSH)), and proteins (reduced BSA) with high sensitivity having a limit of detection of 15.3 nM, 6.0 nM, and 9.2 ng/mL, respectively. Importantly, thiol sensing was selective against analogous small molecules, biomolecules, and proteins devoid of thiol moieties. Cellular imaging demonstrated selective diagnosis of cancer cells by NDI-mal FONPs through quenching of its emission upon interaction with thiols in B16F10 cells due to the high abundance of GSH in cancer cells compared to NIH3T3 cells. NDI-mal FONPs emitted their native fluorescence inside cells subjected to reactive oxygen species mediated thiol oxidation via Fenton's reaction. Notably, GSH-maleimide adduct formation by NDI-mal FONPs displayed notable therapeutic efficacy against cancer cells having â¼2.4-fold higher killing of B16F10 in comparison to NIH3T3 cells possibly through oxidative stress induced apoptosis owing to the depletion in the GSH level. Thus, NDI-mal AIE-gen successfully emerged as a selective and sensitive probe toward thiols through thiol-maleimide click chemistry with therapeutic ability against cancer cells in the absence of systematic intervention.
Assuntos
Dimetil Sulfóxido , Compostos de Sulfidrila , Animais , Camundongos , Células NIH 3T3 , Compostos de Sulfidrila/química , Maleimidas/química , Proteínas , Corantes Fluorescentes/química , ÁguaRESUMO
Aggregation-induced emission (AIE)-based fluorescent organic nanoparticles (FONPs) with distinctive characteristics are emerging as superior sensors due to their facile fabrication, high signal-to-noise ratio, and good biocompatibility. The present article delineates the detection and analysis of the redox behavior of the protein disulfide isomerase (PDI) enzyme by exploitation of the AIE of novel naphthalimide (NI) derivatives having thiol (-SH) and disulfide (-S-S-) moieties. Self-aggregated spherical-shaped organic nanoparticles were prepared by synthesized NI-based amphiphiles (NISH, NISS, NINSS, and TNINSH) through J-type aggregation in DMSO-water (fw = 99 vol %). Naphthyl residue containing NI-derived amphiphiles (NINSS and TNINSH) exhibited AIE (blue and yellow) at 470 and 550 nm, respectively, in DMSO-water (fw = 99 vol %). NINSS and TNINSH FONPs were suitably utilized in sensing PDI through their redox nature of thiol-disulfide exchange. Fluorescence quenching of NINSS FONPs was observed due to reduction of disulfide to thiol by PDI, whereas emission intensity was progressively red-shifted and enhanced ("Dual-AIE") for TNINSH (containing ER-targeting N-tosylethylenediamine), owing to oxidation of thiol to disulfide by PDI. NINSS and TNINSH FONPs were found to be highly efficient in sensing PDI through the AIE-based "fluorescence off/on" mechanism having limits of detection of â¼12.6-17.7 and â¼11.7-16.5 ng/mL, respectively. In vitro cell imaging for NIH3T3 (noncancer) and B16F10 (melanoma) cells with NINSS and TNINSH FONPs displayed excellent diagnosis of eukaryotic cells upon interaction with indigenous PDI. Notably, detection of cancer cells was more sensitive over the noncancerous cells by these FONPs due to overexpression of PDI within cancer cells.
Assuntos
Dimetil Sulfóxido , Isomerases de Dissulfetos de Proteínas , Camundongos , Animais , Células NIH 3T3 , Naftalimidas , Corantes , Dissulfetos , Oxirredução , Compostos de Sulfidrila , ÁguaRESUMO
Glucose oxidase (GOx)-induced cancer starvation has recently emerged for halting the abnormal proliferation of triple-negative breast cancer (TNBC). However, monotherapy with GOx or a conventional chemotherapeutic displays suboptimal efficacy in eliminating tumors and poses impending risks to healthy tissues. To augment therapeutic efficacy and tumor selectivity, folic acid (FA)-functionalized carbon dots (CDs) embedded with GOx and paclitaxel (PTX) [FA-CD-(PTX-GOx)] was developed that showed the efficient killing of TNBC, MDA-MB-468 cells over noncancerous HEK 293 cells through synergistic effects of cancer starvation-induced oxidative stress and chemotherapy. The cargo-laden FA-CD complex resulted in a 4-8 fold increase in cancer cell death at 60 µg/mL when compared to standalone therapy with the native compounds and individually loaded cargo on FA-CD. This improved cancer cell killing efficacy of the FA-CD-(PTX-GOx) complex could be endorsed by folate receptor (FR)-mediated target-specific cellular internalization of the FA-CD complex. The antitumorigenic efficacy of the FA-CD-(PTX-GOx) complex was further validated in a three-dimensional (3D) breast tumor spheroid model. A significant 4.5-fold reduction in spheroid dimension along with antiproliferation was observed with time up to 72 h following exposure to the FA-CD-(PTX-GOx) complex. This antitumorigenic potential of FA-CD-(PTX-GOx) could be attributed to the enhanced intratumoral reactive oxygen species generation following glucose depletion by GOx that has been facilitated by the chemotherapeutic efficacy of PTX resulting in the efficient killing of cancer cells. The present study provides a novel strategy of FR-mediated fluorescent CD-enabled combined formulation of GOx and PTX for the target-specific superior killing of TNBC cells in the synergism of glucose starvation with chemotherapy.
Assuntos
Paclitaxel , Neoplasias de Mama Triplo Negativas , Carbono/farmacologia , Linhagem Celular Tumoral , Ácido Fólico/farmacologia , Glucose , Glucose Oxidase , Células HEK293 , Humanos , Paclitaxel/farmacologia , Neoplasias de Mama Triplo Negativas/tratamento farmacológicoRESUMO
This article demonstrates target-specific cellular imaging of GABA (γ-aminobutyric acid) receptor (GABAAR)-enriched cells (SH-SY5Y and A549) with therapeutic efficacy by naphthalene diimide (NDI)-derived fluorescent organic nanoparticles (FONPs). Self-assembly-driven formation of spherical organic particles by nipecotic-acid-tethered l-aspartic acid appended NDI derivative (NDI-nip) took place in DMSO-water through J-type aggregation. NDI-nip having a naphthyl residue and a nipecotic acid unit at both terminals exhibited aggregation-induced emission (AIE) at and above 60% water content in DMSO because of excimer formation at λem = 579 nm. The orange-emitting NDI-nip FONPs (1:99 v/v DMSO-water) having excellent cell viability and high photostability were used for selective bioimaging and killing of GABAAR-overexpressed cancer cells through target-specific delivery of the anticancer drug curcumin. The fluorescence intensity of NDI-nip FONPs were quenched in GABAAR-enriched neuroblastoma cells (SH-SY5Y) and cancerous cells (A549). Notably, in the presence of GABA, the NDI-nip FONPs exhibited their native fluorescence within the same cell lines. Importantly, no such quenching and regaining of NDI-nip FONP emission in the presence of GABA was noted in the case of the noncancerous cell NIH3T3. The killing efficiency of curcumin-loaded NDI-nip FONPs ([curcumin] = 100 µM and [NDI-nip FONPs] = 50 µM) was significantly higher in the cases of SH-SY5Y (88 ± 3%) and A549 (72 ± 2%) than in NIH3T3 (37 ± 2). The presence of a nipecotic acid moiety facilitated the selective cellular internalization of NDI-nip FONPs into GABAAR-overexpressing cells. Hence, these orange-emitting NDI-nip FONPs may be exploited as a targeted diagnostic probe as well as a drug delivery vehicle for GABAAR-enriched cancer cells.
Assuntos
Curcumina , Nanopartículas , Neoplasias , Ácidos Nipecóticos , Receptores de GABA-A , Células A549 , Animais , Linhagem Celular Tumoral , Curcumina/farmacologia , Dimetil Sulfóxido , Excipientes , Humanos , Imidas , Camundongos , Células NIH 3T3 , Nanopartículas/química , Naftalenos , ÁguaRESUMO
Mucormycosis due to Mucorales is reported at large numbers in uncontrolled diabetics across India, but systematic multicenter epidemiological study has not been published yet. The present prospective study was conducted at four major tertiary care centers of India (two in north and two in south India) during 2013-2015 to compare the epidemiology, treatment strategies and outcome of mucormycosis between the two regions. Molecular techniques were employed to confirm the identity of the isolates or to identify the agent in biopsy samples. A total of 388 proven/probable mucormycosis cases were reported during the study period with overall mortality at 46.7%. Uncontrolled diabetes (n = 172, 56.8%) and trauma (n = 31, 10.2%) were the common risk factors. Overall, Rhizopus arrhizus (n = 124, 51.9%) was the predominant agent identified, followed by Rhizopus microsporus (n = 30, 12.6%), Apophysomyces variabilis (n = 22, 9.2%) and Rhizopus homothallicus (n = 6, 2.5%). On multivariate analysis, the mortality was significantly associated with gastrointestinal (OR: 18.70, P = .005) and pulmonary infections (OR: 3.03, P = .015). While comparing the two regions, majority (82.7%) cases were recorded from north India; uncontrolled diabetes (n = 157, P = .0001) and post-tubercular mucormycosis (n = 21, P = .006) were significantly associated with north Indian cases. No significant difference was noted among the species of Mucorales identified and treatment strategies between the two regions. The mortality rate was significantly higher in north Indian patients (50.5%) compared to 32.1% in south India (P = .016). The study highlights higher number of mucormycosis cases in uncontrolled diabetics of north India and emergence of R. microsporus and R. homothallicus across India causing the disease.
Assuntos
Gerenciamento Clínico , Mucormicose/diagnóstico , Mucormicose/epidemiologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Criança , Pré-Escolar , Complicações do Diabetes , Feminino , Humanos , Índia/epidemiologia , Lactente , Masculino , Pessoa de Meia-Idade , Técnicas de Diagnóstico Molecular , Mucorales/classificação , Mucorales/genética , Mucorales/isolamento & purificação , Mucormicose/mortalidade , Mucormicose/terapia , Estudos Prospectivos , Fatores de Risco , Análise de Sobrevida , Centros de Atenção Terciária , Resultado do Tratamento , Ferimentos e Lesões/complicações , Adulto JovemRESUMO
BACKGROUND: Alzheimer's disease (AD), a common protein misfolding progressive neurodegenerative disorder, is one of the most common forms of dementia. Amyloid precursor protein (APP) derived amyloid-ß (Aß) protein accumulate into interneuronal spaces and plays a crucial role in the disease progression and its pathology. The aggregated Aß exerts its neurotoxic effects by inducing apoptosis and oxidative damage in neuronal cells. OBJECTIVES: We have investigated the effects of a synthesized Pro-Drug peptide (PDp) on Aß1-40 induced cytotoxicity in human neuroblastoma SH-SY5Y cells, represents one of the most effective strategies in combating human AD. METHODS: Cells were treated with Aß1-40 to induce cytotoxicity in the experimental model of AD to screen the inhibitory effect of PDp. Assays for cell viability, reactive oxygen species (ROS) generation, levels of intracellular free Ca2+ and expression of key apoptotic proteins were assessed by Western Blotting. RESULTS: Our results showed that Aß1-40 induces for 24h caused reduce cell viability, imbalance in Ca2+ homeostasis and increase in neuronal apoptosis in vitro. Treatment with PDp could effectively ameliorated Aß1-40 induced neurotoxicity and attenuates ROS generation that mediates apoptotic signaling through Bcl-2, Bax, Caspase-3 activity and cytochrome c in the cells. CONCLUSION: These findings suggested that PDp has potential role as a neuroprotective and therapeutic agent for combating human AD.
Assuntos
Peptídeos beta-Amiloides/toxicidade , Apoptose/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Fragmentos de Peptídeos/toxicidade , Peptídeos beta-Amiloides/farmacologia , Análise de Variância , Cálcio/metabolismo , Caspase 3/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Citocromos c/metabolismo , Humanos , Líquido Intracelular/efeitos dos fármacos , Líquido Intracelular/metabolismo , L-Lactato Desidrogenase/metabolismo , Neuroblastoma/patologia , Fragmentos de Peptídeos/farmacologia , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Proteína X Associada a bcl-2/metabolismoRESUMO
Alzheimer's disease is most common neurodegenerative disorder and is characterized by increased production of soluble amyloid-ß oligomers, the main toxic species predominantly formed from aggregation of monomeric amyloid-ß (Aß). Increased production of Aß invokes a cascade of oxidative damages to neurons and eventually leads to neuronal death. This study was aimed to investigate the neuroprotective effects of a ß-sheet breaker α/ß-hybrid peptide (BSBHp) and the underlying mechanisms against Aß40 -induced neurotoxicity in human neuroblastoma SH-SY5Y cells. Cells were pretreated with the peptide Aß40 to induce neurotoxicity. Assays for cell viability, cell membrane damage, cellular apoptosis, generation of reactive oxygen species (ROS), intracellular free Ca2+ , and key apoptotic protein levels were performed in vitro. Our results showed that pretreatment with BSBHp significantly attenuates Aß40 -induced toxicity by retaining cell viability, suppressing generation of ROS, Ca2+ levels, and effectively protects neuronal apoptosis by suppressing pro-apoptotic protein Bax and up-regulating antiapoptotic protein Bcl-2. These results suggest that α/ß-hybrid peptide has neuroprotective effects against Aß40 -induced oxidative stress, which might be a potential therapeutic agent for treating or preventing neurodegenerative diseases.
Assuntos
Peptídeos beta-Amiloides/metabolismo , Apoptose/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Peptídeos/farmacologia , Western Blotting , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Genes bcl-2/genética , Humanos , Microscopia Eletrônica de Transmissão , Fármacos Neuroprotetores/química , Peptídeos/química , Proteína X Associada a bcl-2/genéticaRESUMO
Combined effects of alprazolam (Alp), a member of benzodiazepine group of drugs and caffeine on human cell lines, HeLa and THP1 were investigated in this study. Alp mediated cytotoxicity was enhanced while caffeine was present. The cell death was confirmed by observing morphological changes, LDH assay and membrane anisotropic study. Also such combined effects induced elevated level of ROS and depletion of GSH. The mechanism of cell death induced by simultaneous treatment of Alp and caffeine was associated with the calcium-mediated activation of mu-calpain, release of lysosomal protease cathepsin B, activation of PARP and cleavage of caspase 3. Our results indicate that, Alp alone induces apoptosis in human cells but in the presence of caffeine it augments necrosis in a well-regulated pathway. Thus our observations strongly suggest that, alprazolam and caffeine together produce severe cytotoxicity in human cell lines.