Auditory neuropathy spectrum disorder (ANSD) represents a variety of sensorineural deafness conditions characterized by abnormal inner hair cells and/or auditory nerve function, but with the preservation of outer hair cell function. ANSD represents up to 15% of individuals with hearing impairments. Through mutation screening, bioinformatic analysis and expression studies, we have previously identified several apoptosis-inducing factor (AIF) mitochondria-associated 1 (AIFM1) variants in ANSD families and in some other sporadic cases. Here, to elucidate the pathogenic mechanisms underlying each AIFM1 variant, we generated AIF-null cells using the clustered regularly interspersed short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) system and constructed AIF-wild type (WT) and AIF-mutant (mut) (p.‍T260A, p.‍R422W, and p.‍R451Q) stable transfection cell lines. We then analyzed AIF structure, coenzyme-binding affinity, apoptosis, and other aspects. Results revealed that these variants resulted in impaired dimerization, compromising AIF function. The reduction reaction of AIF variants had proceeded slower than that of AIF-WT. The average levels of AIF dimerization in AIF variant cells were only 34.5%‍‒‍49.7% of that of AIF-WT cells, resulting in caspase-independent apoptosis. The average percentage of apoptotic cells in the variants was 12.3%‍‒‍17.9%, which was significantly higher than that (6.9%‍‒‍7.4%) in controls. However, nicotinamide adenine dinucleotide (NADH) treatment promoted the reduction of apoptosis by rescuing AIF dimerization in AIF variant cells. Our findings show that the impairment of AIF dimerization by AIFM1 variants causes apoptosis contributing to ANSD, and introduce NADH as a potential drug for ANSD treatment. Our results help elucidate the mechanisms of ANSD and may lead to the provision of novel therapies.
Humans ; Apoptosis Inducing Factor/metabolism* ; NAD/metabolism* ; Dimerization ; Apoptosis

OBJECTIVE: To investigate the effect of lycium barbarum polysaccharide (LBP) alone or combined with tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) on the apoptosis of leukemia cell lines with MLL gene-rearrangement, and to explore the cell apoptotic pathway after the combined action. METHODS: MLL-ALL cell line KOCL44 and KOCL45 were selected as the research object, then the control and experimental groups were set up. The cell survival rate was measured by the trypan blue dye exclusion method, the cell early apoptosis and expression of death receptors on the cell surface were detected by flow cytometry with Annexin-V/PI double staining. The protein level of caspase-8, BID, caspase-3, caspase-9, BAD, BCL-2, as well as mitochondrial and cytosol Cyto-C were detected by Western blot. RESULTS: LBF combined with TRAIL inhibited the growth of KOCL44 and KOCL-45 cells and showed the synergistic effect, the results of flow cytometry with Amnexiu V/PI double staining were consistent with above-mentioned results. After treatment of KOCL44 and KOCL45 cells with LBF plus TRAIL, the significant expression of DR4 on cell surface was not found, while the expression of DR4 receptor was enhanced significantly, the pro-apoptotic proteins including caspase-8, BID, caspase-3, caspase-9 and BAD were activated significantly and BCL-2 was suppressed significantly with time-dependent manner. The expression of mitochondria cyto-C in KOCL44 and KOCL45 decreased along with prolonging of treatment time (r=-0.95, r=-0.866), while the expression of cytosol cyto-C in KOCL44 and KOCL45 increased along with prolonging of treatment time (r=0.883, r=0.903). CONCLUSION The combination of LBP and TRAIL significantly increases the apoptosis of KOCL44 and KOCL45, and the LBP and TRAIL can up-regulate the expression of TRAIL death receptor-DR5 on the cell surface, activate the pathway of caspase and mito-chrondia mitachondria, thus enhance the sensitivity of KOCL44 and KOCL45 to TRAIL induced apoptosis through both mitochondrial and apoptotic pathway.
Apoptosis ; Caspase 8 ; Drugs, Chinese Herbal ; Leukemia, Myeloid, Acute ; Receptors, TNF-Related Apoptosis-Inducing Ligand ; TNF-Related Apoptosis-Inducing Ligand ; Tumor Necrosis Factor-alpha

Inflammation is the crucial biological process of immune system which acts as body's defense and protective response against the injuries or infection. However, the systemic inflammation devotes the adverse effects such as multiple inflammation associated diseases. One of the best ways to treat this entity is by blocking the tumor necrosis factor alpha (TNF-α) and TNF-related apoptosis-inducing ligand (TRAIL) to avoid the proinflammation cytokines production. Thus, this study aims to evaluate the potency of Sambucus bioactive compounds as anti-inflammation through in silico approach. In order to assess that, molecular docking was performed to evaluate the interaction properties between the TNF-α or TRAIL with the ligands. The 2D structure of ligands were retrieved online via PubChem and the 3D protein modeling was done by using SWISS Model. The prediction results of the study showed that caffeic acid (−6.4 kcal/mol) and homovanillic acid (−6.6 kcal/mol) have the greatest binding affinity against the TNF-α and TRAIL respectively. This evidence suggests that caffeic acid and homovanillic acid may potent as anti-inflammatory agent against the inflammation associated diseases. Finally, this study needs further examination and evaluation to validate the potency of Sambucus bioactive compounds.
Biological Processes ; Computational Biology ; Computer Simulation ; Cytokines ; Homovanillic Acid ; Immune System ; Inflammation ; Ligands ; Plants ; Sambucus ; TNF-Related Apoptosis-Inducing Ligand ; Tumor Necrosis Factor-alpha

PURPOSE: This study was conducted to verify the induction and mechanism of selective apoptosis in G361 melanoma cells using anti-HER2 antibody-conjugated gold nanoparticles (GNP-HER2). MATERIALS AND METHODS: Following GNP-HER2 treatment of G361 cells, cell cycle arrest and apoptosis were measured by WST-1 assay, Hemacolor staining, Hoechst staining, immunofluorescence staining, fluorescence-activated cell sorting analysis, and Western blotting.
Actins ; Apoptosis Inducing Factor ; Apoptosis ; Blotting, Western ; Caspase 3 ; Caspases ; Cell Adhesion ; Cell Cycle ; Cell Cycle Checkpoints ; Cell Death ; Cyclin A ; Cyclin D1 ; Cyclin E ; Cyclins ; Cytochromes c ; Cytoplasm ; DNA Fragmentation ; Down-Regulation ; Flow Cytometry ; Fluorescent Antibody Technique ; Focal Adhesions ; Melanoma ; Mitochondria ; Nanoparticles ; Phosphotransferases ; Receptor, Epidermal Growth Factor ; Up-Regulation

Psoriasis is a chronic, recurrent, heterogeneous, cutaneous inflammatory skin disease for which there is no cure. It affects approximately 7.5 million people in the United States. Currently, several biologic agents that target different molecules implicated in the pathogenic processes of psoriasis are being assessed in diverse clinical studies. However, relapse usually occurs within weeks or months, meaning there is currently no cure for psoriasis. Therefore, recent studies have discovered diverse new potential treatments for psoriasis: inhibitors of bacteria such as Staphylococcus aureus, tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and neuropilin 1 (NRP1). A promising approach that has recently been described involves modifying antimicrobial peptides to develop new cutaneous anti-bacterial agents that target inflammatory skin disease induced by Staphylococcus. Increased expression of TRAIL and its death receptors DR4 and DR5 has been implicated in the pathogenesis of plaque psoriasis. In addition, TRAIL has the ability to inhibit angiogenesis by inducing endothelial cell death and by negative regulation of VEGF-induced angiogenesis via caspase-8-mediated enzymatic and non-enzymatic functions. Since NRP1 regulates angiogenesis induced by multiple signals, including VEGF, ECM and semaphorins, and also initiates proliferation of keratinocytes through NF-κB signaling pathway in involved psoriatic skin, targeting NRP1 pathways may offer numerous windows for intervention in psoriasis. In this review, we will focus on the current knowledge about the emerging role of synthetic antimicrobial peptides, TRAIL and NRP1 blocking peptides in the pathogenesis and treatment of psoriasis.
Anti-Bacterial Agents ; Bacteria ; Biological Factors ; Endothelial Cells ; Keratinocytes ; Necrosis ; Neuropilin-1 ; Peptides ; Psoriasis ; Receptors, Death Domain ; Recurrence ; Semaphorins ; Skin ; Skin Diseases ; Staphylococcus ; Staphylococcus aureus ; Therapeutic Uses ; TNF-Related Apoptosis-Inducing Ligand ; United States ; Vascular Endothelial Growth Factor A

OBJECTIVES: To investigate whether the cytotoxic effect of Cimicifuga rhizoma extract is associated with cell death in the human keratinocyte (HaCaT) and human melanoma cell lines (G361). METHODS: Apoptosis induced by Cimicifuga rhizoma extract was confirmed by water-soluble tetrazolium salts-1 (WST-1) assay, immunocytochemistry, and western blot. Additionally, the release of cytochrome c and apoptosis-inducing factor (AIF) was visualized by confocal laser scanning microscopy. RESULTS: The results showed that Cimicifuga rhizoma extract significantly reduced the viability of G361 cells with half-maximal inhibitory concentration (IC 50) of 200 µg/ml, and the apoptotic process was found to occur via the activation of caspase-3 and caspase-9 pathways. Besides, the release of cytochrome c and AIF was also detected. CONCLUSIONS: This study suggests that Cimicifuga rhizoma extract causes apoptosis of human melanoma cells through the intrinsic apoptotic pathway.
Apoptosis Inducing Factor ; Apoptosis ; Blotting, Western ; Caspase 3 ; Caspase 9 ; Cell Death ; Cell Line ; Cimicifuga ; Cytochromes c ; Humans ; Immunohistochemistry ; Keratinocytes ; Melanoma ; Microscopy, Confocal

OBJECTIVE: Patients with bipolar disorder (BD) exhibit peripheral low-grade inflammation. The aim of the current study was to investigate the involvement of hitherto unexplored components of the tumor necrosis factor (TNF) superfamily in BD. METHODS: Eighty patients with type I BD and 50 healthy controls matched for age and gender were enrolled in this study. All subjects were assessed with the Mini-Plus to evaluate psychiatric comorbidities; the Young Mania Rating Scale and the Hamilton Depression Rating Scale to evaluate manic and depressive symptoms severity, respectively. TNF superfamily molecules (TNF, TNF-related weak inducer of apoptosis [TWEAK], TNF-related apoptosis-inducing ligand [TRAIL], soluble TNF receptor type 1 [sTNFR1], and soluble TNF receptor type 2 [sTNFR2]) levels were measured by ELISA. RESULTS: Patients with BD, regardless of mood state, presented increased plasma levels of sTNFR1 and TWEAK in comparison with controls. CONCLUSION: These findings corroborate the view that TNF superfamily may play a role in BD pathophysiology.
Apoptosis ; Bipolar Disorder* ; Comorbidity ; Depression ; Enzyme-Linked Immunosorbent Assay ; Humans ; Inflammation ; Plasma* ; Receptors, Tumor Necrosis Factor ; TNF-Related Apoptosis-Inducing Ligand ; Tumor Necrosis Factor-alpha*

In keloids, the mechanism underlying the excessive accumulation of extracellular matrix after injury of the skin is unclear, and there is no effective treatment because of the incomplete understanding of their pathogenesis; thus, a high recurrence rate is observed. We studied a new marker of keloids to determine a new treatment strategy. First, the keloid gene expression profile (GSE44270) was analyzed (downloaded from the Gene Expression Omnibus database) and the new keloid marker candidate, epidermal growth factor (EGF)-like repeats and discoidin I-like domains 3 (EDIL3) which were upregulated in keloid samples was identified. Knockdown of EDIL3 is known to suppresses angiogenesis by downregulating relevant inhibitory factors that can limit the supply of survival factors to tumor cells from the circulation via the vascular endothelial cells. In keloids, the mechanism of action of EDIL3 may be similar to that in tumors; the inhibition of apoptosis in tumor cells via a reduction in the apoptosis of blood vessels by upregulating an angiogenic factor. To determine whether EDIL3 is involved in keloid formation, we performed knockdown of EDIL3 in keloid fibroblasts in vitro by transfection with anti-EDIL3 small interfering RNA (via microporation). EDIL3 was upregulated in keloid fibroblasts compared with normal fibroblasts in collagen type I, II and III. Our results indicate the control of EDIL3 expression may be a new promising treatment of keloid disease also the molecular targeting of EDIL3 may improve the quality of treatment and reduce the formation of keloids.
Angiogenesis Inducing Agents ; Apoptosis ; Blood Vessels ; Cicatrix* ; Collagen ; Collagen Type I ; Endothelial Cells ; Epidermal Growth Factor* ; Extracellular Matrix ; Fibroblasts ; Gene Expression ; In Vitro Techniques ; Keloid* ; Recurrence ; RNA, Small Interfering ; Skin ; Transcriptome ; Transfection

The purpose of this study was to investigate the effect of inhibition of calpain on retinal ganglion cell-5 (RGC-5) necroptosis following oxygen glucose deprivation (OGD). RGC-5 cells were cultured in Dulbecco's-modified essential medium and necroptosis was induced by 8-h OGD. PI staining and flow cytometry were performed to detect RGC-5 necrosis. The calpain expression was detected by Western blotting and immunofluorescence staining. The calpain activity was tested by activity detection kit. Flow cytometry was used to detect the effect of calpain on RGC-5 necroptosis following OGD with or without N-acetyl-leucyl-leucyl-norleucinal (ALLN) pre-treatment. Western blot was used to detect the protein level of truncated apoptosis inducing factor (tAIF) in RGC-5 cells following OGD. The results showed that there was an up-regulation of the calpain expression and activity following OGD. Upon adding ALLN, the calpain activity was inhibited and tAIF was reduced following OGD along with the decreased number of RGC-5 necroptosis. In conclusion, calpain was involved in OGD-induced RGC-5 necroptosis with the increased expression of its downstream molecule tAIF.
Animals ; Apoptosis Inducing Factor ; biosynthesis ; genetics ; Calpain ; biosynthesis ; genetics ; Gene Expression Regulation ; drug effects ; Glucose ; metabolism ; Humans ; Leupeptins ; administration & dosage ; Mice ; Oxygen ; metabolism ; Retinal Ganglion Cells ; metabolism ; pathology ; Retinal Necrosis Syndrome, Acute ; genetics ; pathology

Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is a member of the TNF super family found in recent years, which widely exists in the body tissues and participates in the immune regulation, immune stability, and immune surveillance of the human body. The TRAIL receptor is expressed in the surface of a variety of cells. Recent studies show that TRAIL induces the apoptosis of tumor cells and has no significant toxic effect on normal cells. Its anti-tumor activity and safety have been widely recognized. The development of prostate cancer is regulated by the mechanisms of cell apoptosis. TRAIL can induce the apoptosis of prostate cancer cells, and therefore has a great application value in the treatment of prostate cancer.
Antineoplastic Agents ; therapeutic use ; Apoptosis ; Apoptosis Regulatory Proteins ; Humans ; Male ; Membrane Glycoproteins ; Prostatic Neoplasms ; drug therapy ; pathology ; Receptors, TNF-Related Apoptosis-Inducing Ligand ; physiology ; therapeutic use ; TNF-Related Apoptosis-Inducing Ligand ; Tumor Necrosis Factor-alpha