Table 1.

Characteristics of apoptosis, autophagy and necrosis pathways

	Morphological and biochemical features and modulators of cell death	Methods of detection
Apoptosis	Morphological features: cellular shrinking, condensation and margination of chromatin, nuclear fragmentation and DNA laddering, plasma membrane budding and formation of apoptotic bodies in cytoplasm. Not surrounded by tissue injury of inflammation. Biochemical features: caspase-dependent cell death pathway. Activation: activation of intrinsic apoptotic pathway; BCL-2, c-FLIP, survivin IAP–antisense mRNA technology; recombinant TRAIL for DR4 and/or DR5 receptor; E2F-1 gene therapy; TWEAK (tumor necrosis factor-related weak inducer of apoptosis) is a cytokine belonging to TNF-ligand family for Tweak-receptor inducing apoptosis. Inhibition: natural and synthetic inhibitors of caspases; nitrosylation of caspase 9 or 3; c-Jun–mRNA antisense technology; CEP 1347–inhibitor of JNK signaling blocks Aβ-induced cortical neuron apoptosis.	Microscopic techniques: cellular features by light microscopy, nuclear DNA analysis by fluorescent stains (annexin V), confocal laser microscopy and electron microscopy. Assessment of DNA fragmentation: enzyme-linked immunosorbent assay, terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling assay, comet method, DNA diffusion, immunohistochemistry for single-stranded DNA and gel electrophoresis. Flow cytometry: cell cycle. Laser scanning cytometry: DNA content, phosphatidylserine translocation, inner mitochondrial transmembrane potential and caspase activity. Gene expression: northern blot, RNA protection assay, reverse transcription–polymerase chain reaction and immunohistochemistry. Evaluation of apoptosis-associated proteins: enzyme-linked immunosorbent assay, western blot and electrophoretic mobility shift assay.
Autophagy	Morphological features: partial chromatin condensation, no DNA laddering, cell membrane blebbing and formation of more autophagosome. Biochemical features: caspase-independent cell death pathway. Activators: conventional cytotoxic drugs and irradiation; BCR-ABL tyrosine kinase inhibitor–imatinib; anti-EGFR–cetuximab; proteasome inhibitors; TRAIL and histone deacetylase inhibitors; mTOR inhibitors and its analogs; ATP-competitive inhibitors of mTORC1 and mTORC2; dual PI3K-mTOR inhibitor; antidiabetic drug–metformin; serotonin reuptake inhibitor–fluoxetine; norepinephrine reuptake inhibitor–maprotiline; antiepileptic drug–valproic. Inhibitors: antibody against EGFR–cetuximab; Class III PI3K inhibitors-3-methyadenine, wortmannin and LY294002; antimalarial drugs–hydroxychloroquine; vacuolar ATPase–bafilomycin A1; lysosomotropic drug–monensin; microtubule-disrupting agents–taxanes, nocodazole, colchicine, vinca alkaloids; antidepressant drug–clomipramine; antischistome agent–lucanthone (autophagosome degradation) .	Electron microscopy, immunohistochemical staining of microtubule-associated protein 1 light chain 3 (LC3) as a general marker for autophagic membranes, monodansylcadaverine staining of autophagic vacuoles and protein degradation assays.
Necrosis	Morphological features: cell size increases, clumping and random degradation of nuclear DNA, cell membrane swelling and rupture, swelling of organelles, gain in cell volume (oncosis), organelle degeneration mitochondrial swelling and increased vacuolation. Activation: hyperactivation of poly(ADP-ribose) polymerase 1 (PARP1) enzyme with depletion of β-nicotinamide adenine dinucleotide and of ATP, hypoxic injury and oxidative stress (ROS/reactive nitrogen species); chetomin–inhibitor of tumor growth by inducing necrosis in vivo; dimethoxy-naphthoquinone–generation of ROS and induces apoptosis or necrosis; myristoleic acid methyl ester–induces apoptosis and necrosis in prostate cancer cells; sterigmatocystin–a mycotoxin inhibits DNA synthesis and causes necrosis. Inhibition: necrox-2 [5-(1,1-dioxo-thiomorpholin-4-ylmethyl)-2-phenyl-1H-indol-7-yl]-(1-methanesulfonyl-piperidin-4-yl)-amine] and necrox-5 [5-(1,1-dioxo-thiomorpholin-4-ylmethyl)-2-phenyl-1H-indol-7-yl]-(tetrahydro-pyran-4-ylmethyl)-amine]–is a cell-permeable necrosis inhibitor selectively locks oxidative stress-induced necrosis with antioxidant property; tyrphostin AG 126–reduces LPS-induced tyrosine phosphorylation of p42MAPK; cyclosporin A–inhibitor of the mitochondrial permeability transition pore (MPTP) and prevents necrosis; IM-54 (indolylmaleimide derivative)–inhibits necrotic cell death induced by H2O2 in promyelocytic leukemia HL-60 cells; PARP inhibitor VIII, PJ34 [2-(dimethylamino)-N-(6-oxo-5,6-dihydrophenanthridin-2-yl)acetamide hydrochloride]–inhibitor of PARP-1 and PARP-2 and inhibits necrosis; IM-54 [2-(1H-indol-3-yl)-3-pentylamino-maleimide]—a selective inhibitor of oxidative stress-induced necrosis.	Electron microscopy; nuclear negative staining; ethidium homodimer III DNA assay; detection of inflammation and damage in surrounding tissues.