Mitochondrial Apoptosis Signaling Pathway Research Articles

Exploration of mechanism of total triterpenoids from fruits of Chaenomeles speciosa against senescent GES-1 cells induced by D-galactose based on Gln/GLS1/α-KG metabolic axis and mitochondrial apoptosis signaling pathway

Total triterpenoids from the fruits of Chaenomeles speciosa(TCS) are active components in the prevention and treatment of gastric mucosal damage, which have potential anti-aging effects. However, it is still unclear whether TCS can improve gastric aging, especially its molecular mechanism against gastric aging. On this basis, this study explored the effect and mechanism of TCS on senescent GES-1 cells induced by D-galactose(D-gal) to provide scientific data for the clinical use of TCS to prevent gastric aging. GES-1 cells cultured in vitro and those transfected with overexpression GLS1(GLS1-OE) plasmid of glutaminase 1(GLS1) were induced to aging by D-gal, and then TCS and or GLS1 inhibitor bis-2-(5-phenylacetamido-1,3,4-thiadiazol-2-yl) ethyl sulfide(BPTES) were given. Cell survival rate, positive rate of β-galactosidase(SA-β-gal) staining, mitochondrial membrane potential(MMP), and apoptosis were investigated. GLS1 activity, levels of glutamine(Gln), glutamate(Glu), α-ketoglutarate(α-KG), urea, and ammonia in supernatant and cells were detected by enzyme-linked immunosorbent assay(ELISA) and colorimetric methods. The mRNA and protein expressions of GLS1 and the related genes of the mitochondrial apoptosis signaling pathway were measured by real-time fluorescence quantitative PCR and Western blot. The results manifested that compared with the D-gal model group and GLS1-OE D-gal model group, TCS significantly decreased the SA-β-gal staining positive cell rate and MMP of D-gal-induced senescent GES-1 cells and GLS1-OE senescent GES-1 cells, inhibited the survival of senescent cells, and promoted their apoptosis(P<0.01). It decreased the activity of GLS1 and the content of Gln, Glu, α-KG, urea, and ammonia in supernatant and cell(P<0.01), reduced the concentration of cytochrome C(Cyto C) in mitochondria and the mRNA and protein expressions of GLS1 and proliferating nuclear antigen in cells(P<0.01). The mRNA expression of Bcl-2 and Bcl-xl, the protein expression of pro-caspase-9 and pro-caspase-3, and the ratio of Bcl-2/Bax and Bcl-xl/Bad in cells were decreased(P<0.01). Cyto C concentration in the cytoplasm, the mRNA expressions of Bax, Bad, apoptosis protease activating factor 1(Apaf-1), and protein expressions of cleaved-caspase-9, cleaved-caspase-3, cleaved-PARP-1 were increased(P<0.01). The aforementioned results indicate that TCS can counteract the senescent GES-1 cells induced by D-gal, and its mechanism may be closely related to suppressing the Gln/GLS1/α-KG metabolic axis, activating the mitochondrial apoptosis pathway, and thereby accelerating the apoptosis of the senescent cells and eliminating senescent cells.

Research progress on the role of mitochondria in the process of hepatic ischemia-reperfusion injury.

During liver ischemia-reperfusion injury, existing mechanisms involved oxidative stress, calcium overload, and the activation of inflammatory responses involve mitochondrial injury. Mitochondrial autophagy, a process that maintains the normal physiological activity of mitochondria, promotes cellular metabolism, improves cellular function, and facilitates organelle renewal. Mitochondrial autophagy is involved in oxidative stress and apoptosis, of which the PINK1-Parkin pathway is a major regulatory pathway, and the deletion of PINK1 and Parkin increases mitochondrial damage, reactive oxygen species production, and inflammatory response, playing an important role in mitochondrial quality regulation. In addition, proper mitochondrial permeability translational cycle regulation can help maintain mitochondrial stability and mitigate hepatocyte death during ischemia-reperfusion injury. This mechanism is also closely related to oxidative stress, calcium overload, and the aforementioned autophagy pathway, all of which leads to the augmentation of the mitochondrial membrane permeability transition pore opening and cause apoptosis. Moreover, the release of mitochondrial DNA (mtDNA) due to oxidative stress further aggravates mitochondrial function impairment. Mitochondrial fission and fusion are non-negligible processes required to maintain the dynamic renewal of mitochondria and are essential to the dynamic stability of these organelles. The Bcl-2 protein family also plays an important regulatory role in the mitochondrial apoptosis signaling pathway. A series of complex mechanisms work together to cause hepatic ischemia-reperfusion injury (HIRI). This article reviews the role of mitochondria in HIRI, hoping to provide new therapeutic clues for alleviating HIRI in clinical practice.

Effects of ginseng peptides on the hypoglycemic activity and gut microbiota of a type 2 diabetes mellitus mice model

Ginseng peptide (GP) (100, 200, 400 mg/kg bw) lowered blood glucose levels, explored the relationship between GP and gut microbiota-based hypoglycemic effects. GP significantly reduced fasting blood glucose levels (25.38–13.37 mM), improved insulin resistance (IR) (9.78–5.02), considerably increased liver glycogen content (64.71–97.95 mg/g), glycolytic and gluconeogenesis enzyme activities in T2DM mice, significantly reduced free fatty acids (1.65–0.93 mM), triglycerides (1.55–0.66 mM), total cholesterol (14.75–5.43 mM), and low-density lipoprotein (4.12–2.36 mM) in the serum, significantly increased high-density lipoprotein levels (6.20–17.44 mM) to improve lipid metabolism disorders. According to colonic 16S rDNA sequencing results, GP-H regulated gut microbiota disorders and short chain fatty acid metabolism. Western blot analysis showed that GP could regulate glycogen synthesis, gluconeogenesis, and glucose transport by activating IRS-1/PI3K/Akt and AMPK signaling pathways and improve IR by inhibiting the mitochondrial apoptosis signaling pathway.

Controlled synthesis of zinc oxide nanoparticles through flame spray pyrolysis and evaluation of their anticancer effects against gastric cancer cell

Although zinc oxide nanoparticles (ZnO NPs) have demonstrated promising anticancer activity, their therapeutic performances are frequently influenced by the synthesis routes and associated distinct physicochemical characteristics. Herein, ZnO NPs were synthesized through a cost-effective flame spray pyrolysis-assisted approach and their selective anticancer effects and underlying intrinsic apoptosis signaling pathway in human gastric carcinoma cell line (AGS) were examined. Transmission electron microscopy (TEM), Fourier-transform infrared spectroscopy (FT-IR), X-ray diffraction (XRD), and dynamic light scattering (DLS) investigations were used to characterize the synthesized ZnO NPs. Afterwards, AGS and normal fibroblast cells (L-929) were treated with various concentrations of ZnO NPs, and cell proliferation was examined by the MTT assay to determine the IC50 concentration of NPs. Furthermore, the quantification of ROS and apoptosis was done by fluorometer and flow cytometry assays. Additionally, superoxide dismutase (SOD) and catalase (CAT) activity assays were performed to evaluate the oxidative stress-induced cytotoxicity mediated by ZnO NPs against AGS cells. Moreover, caspase-3/-8/-9 activity and mitochondrial membrane potential (MMP) assays were done. Finally, the expression levels of p53, Bax, Bcl-2, caspase-9/-8/-3, and cytochrome c were assessed by quantitative real time PCR and western blot analyses to explore the probable mitochondrial apoptosis signaling pathway induced by ZnO NPs in AGS cells. The results showed that ZnO NPs with an absorption maximum at 350 nm, an average size of 70 nm, a hydrodynamic radius of 92.89 nm, and a zeta potential of −43.13 mV potentially and selectively inhibited the proliferation of AGS cells in comparison with L-929 cells through a remarkable increase in the production of ROS, induction of apoptosis, reduction of SOD and CAT activity, activation of caspase-9/-3, and MMP disruption. Moreover, ZnO NPs triggered up-regulation of p53, Bax, caspase-9/-3, cytochrome c and down-regulation of Bcl-2 at mRNA and protein levels. This study suggests that synthesized ZnO NPs through flame spray pyrolysis can induce selective anticancer effects against gastric cancer cells via intrinsic apoptosis signaling pathway.

Ferric citrate-induced colonic mucosal damage associated with oxidative stress, inflammation responses, apoptosis, and the changes of gut microbial composition

Ferric citrate (FC) has been used as an iron fortifier and nutritional supplement, which is reported to induce colitis in rats, however the underlying mechanism remains to be elucidated. We performed a 16-week study of FC in male healthy C57BL/6 mice (nine-month-old) with oral administration of Ctr (0.9 % NaCl), 1.25 % FC (71 mg/kg/bw), 2.5 % FC (143 mg/kg/bw) and 5 % FC (286 mg/kg/bw). FC-exposure resulted in colon iron accumulation, histological alteration and reduce antioxidant enzyme activities, such as glutathione (GSH), glutathione peroxidase (GSH-Px), superoxide dismutase (SOD) and total antioxidant capacity (T-AOC), together with enhanced lipid peroxidation level, including malondialdehyde (MDA) level and 4-Hydroxynonenal (4-HNE) protein expression. Exposure to FC was associated with upregulated levels of the interleukin (IL)− 6, IL-1β, IL-18, IL-8 and tumor necrosis factor α (TNF-α), while down-regulated levels of IL-4 and IL-10. Exposure to FC was positively associated with the mRNA and protein expressions of cysteine-aspartic proteases (Caspase)− 9, Caspase-3, Bcl-2-associated X protein (Bax), while negatively associated with B-cell lymphoma 2 (Bcl2) in mitochondrial apoptosis signaling pathway. FC-exposure changed the diversity and composition of gut microbes. Additionally, the serum lipopolysaccharide (LPS) contents increased in FC-exposed groups when compared with the control group, while the expression of colonic tight junction proteins (TJPs), such as Claudin-1 and Occludin were decreased. These findings indicate that the colonic mucosal injury induced by FC-exposure are associated with oxidative stress generation, inflammation response and cell apoptosis, as well as the changes in gut microbes diversity and composition.

Cyt-C Mediated Mitochondrial Pathway Plays an Important Role in Oocyte Apoptosis in Ricefield Eel (Monopterus albus).

Apoptosis plays a key role in the effective removal of excessive and defective germ cells, which is essential for sequential hermaphroditism and sex change in vertebrates. The ricefield eel, Monopterus albus is a protogynous hermaphroditic fish that undergoes a sequential sex change from female to male. Previous studies have demonstrated that apoptosis is involved in sex change in M. albus. However, the apoptotic signaling pathway is unclear. In the current study, we explored the underlying mechanism of apoptosis during gonadal development and focused on the role of the mitochondrial apoptosis signaling pathway in sex change in M. albus. Flow cytometry was performed to detect apoptosis in gonads at five sexual stages and ovary tissues exposed to hydrogen peroxide (H2O2) in vitro. Then the expression patterns of key genes and proteins in the mitochondrial pathway, death receptor pathway and endoplasmic reticulum (ER) pathway were examined. The results showed that the apoptosis rate was significantly increased in the early intersexual stage and then decreased with the natural sex change from female to male. Quantitative real-time PCR revealed that bax, tnfr1, and calpain were mainly expressed in the five stages. ELISA demonstrated that the relative content of cytochrome-c (cyt-c) in the mitochondrial pathway was significantly higher than that of caspase8 and caspase12, with a peak in the early intersexual stage, while the levels of caspase8 and caspase12 peaked in the late intersexual stage. Interestingly, the Pearson’s coefficient between cyt-c and the apoptosis rate was 0.705, which suggests that these factors are closely related during the gonadal development of M. albus. Furthermore, the cyt-c signal was found to be increased in the intersexual stage by immunohistochemistry. After incubation with H2O2, the mRNA expression of mitochondrial pathway molecules such as bax, apaf-1, and caspase3 increased in ovary tissues. In conclusion, the present results suggest that the mitochondrial apoptotic pathway may play a more important role than the other apoptotic pathways in sex change in M. albus.

Mitochondria-targeting folic acid-modified nanoplatform based on mesoporous carbon and a bioactive peptide for improved colorectal cancer treatment

Oral colon-targeted drug delivery systems (OCDDs) are designed to deliver the therapeutic agents to colonic disease sites to improve the effectiveness of drug treatment, increase bioavailability, and reduce systemic side effects and are beneficial for the treatment of colorectal cancer (CRC) and inflammatory bowel disease (IBD). However, concerns about the biosafety of OCDDs are increasing, and changes in the physiological environment of the gastrointestinal tract can affect the therapeutic efficacy of the drug. Herein, we report about an orally administered colon-accumulating mitochondria-targeted drug delivery nanoplatform (M27-39@FA-MCNs), which was synthesized using the small peptide, M27-39, and folic acid (FA)-modified mesoporous carbon nanoparticles (FA-MCNs). The phenolic resin polymerized with phloroglucinol and formaldehyde (PF) was used for fabricating MCNs using a one-step soft-template method. Folic acid (FA) can be covalently combined with chitosan-modified MCNs to obtain FA-MCNs. The M27-39@FA-MCNs were stable with a spherical morphology and an average diameter of 129 nm. The cumulative release rate of M27-39@FA-MCNs in the artificial gastric fluid (pH = 1.2) and intestinal fluid (pH = 6.8) for 6 h was 87.77%. This nanoplatform maintains the advantages of both FA and MCNs to improve the bioactivity of M27-39 with high drug accumulation in colorectal tumor tissues and the ease of excretion, thus ameliorating its biosafety and targetability. Furthermore, M27-39@FA-MCNs induced tumor-cell apoptosis and inhibited tumor growth by disrupting mitochondrial energy metabolism and regulating the mitochondrial apoptosis signaling pathway and immune inflammatory response. Thus, such a mitochondria-targeting FA-modified nanoplatform based on mesoporous carbon and a bioactive peptide may provide a precise strategy for CRC treatment. Statement of significanceIn this study, we constructed an orally administered colon-accumulating mitochondria-targeted drug delivery nanoplatform (M27-39@FA-MCNs), which was synthesized using the small peptide (M27-39) and folic acid-modified mesoporous carbon nanoparticles (FA-MCNs). M27-39@FA-MCNs increased the targeting ability of M27-39 toward mitochondria and colon based on the properties of FA-MCNs; they also increased M27-39 accumulation and residence time in colon tumors. Oral administration of M27-39@FA-MCNs remarkably alleviated colorectal cancer (CRC) by targeting tumor cell mitochondria and interfering with the mitochondrial energy metabolism process, and inducing apoptosis related P53/Caspase-3 mitochondrial pathway activation. Therefore, M27-39@FA-MCNs may provide a safe and precise therapeutic strategy for CRC.

DBDPE and ZnO NPs synergistically induce neurotoxicity of SK-N-SH cells and activate mitochondrial apoptosis signaling pathway and Nrf2-mediated antioxidant pathway

Decabromodiphenyl ethane (DBDPE), a new brominated flame retardant, could negatively affect neurobehavior and pose health risks to humans. Humans are also exposed to widely used nanomaterials. This study investigated the combined toxic effects and action types of DBDPE and Zinc oxide nanoparticles (ZnO NPs) on human neuroblastoma SK-N-SH cells and the toxicity mechanisms. DBDPE inhibited the viability of SK-N-SH cells by 21.87% at 25 mg/L. ZnO NPs synergistically exacerbated the toxic effects of DBDPE. DBDPE and ZnO NPs caused excessive ROS production and inhibition of antioxidant enzyme (SOD and GSH) activity in cells, thus causing oxidative cellular damage. Moreover, DBDPE and ZnO NPs caused apoptosis by disrupting mitochondrial kinetic homeostasis, reducing mitochondrial membrane potential (MMP), increasing cytochrome C release and regulating Bax/Bcl-2 and Caspase-3 mRNA and protein expression. DBDPE and ZnO NPs increased the mRNA expression of nuclear factor erythroid 2- related factor (Nrf2) and its downstream genes. The molecular mechanisms revealed that oxidative stress, apoptosis and mitochondrial dysfunction were the critical factors in combined cytotoxicity. The bioinformatics analysis further indicated that co-exposure affected Nrf2 activation, apoptotic factors expression and mitochondrial fusion. The findings enrich the risk perception of neurotoxicity caused by DBDPE and ZnO NPs.

Aconiti lateralis radix praeparata total alkaloids exert anti-RA effects by regulating NF-κB and JAK/STAT signaling pathways and promoting apoptosis.

Aconiti Lateralis Radix Praeparata (“Fuzi” in Chinese) is one of the traditional herbs widely used to intervene rheumatoid arthritis (RA), while Fuzi total alkaloids (FTAs) are the main bioactive components. However, the treatment targets and specific mechanisms of FTAs against RA have not been fully elucidated. The purpose of the present study was to confirm the anti-rheumatism effects of FTAs and reveal its potential molecular mechanisms. In TNF-α-induced MH7A cells model, we found that FTAs showed inhibitory effects on proliferation. While, FTAs significantly decreased the expression levels of IL-1β, IL-6, MMP-1, MMP-3, PGE2, TGF-β, and VEGF. FTAs also enhanced the progress of apoptosis and arrested the cell cycle at G0/G1 phase to prevent excessive cell proliferation. In addition, FTAs inhibited the hyperactivity of NF-κB and JAK/STAT signaling pathways, and regulated the cascade reaction of mitochondrial apoptosis signaling pathway. The results suggested that FTAs exerted anti-inflammatory effects by inhibiting NF-κB and JAK/STAT signaling pathways, promoted apoptosis by stimulating mitochondrial apoptosis signaling pathway, and inhibited cell proliferation by modulating cell cycle progression.

Multistage-Responsive Gene Editing to Sensitize Ion-Interference Enhanced Carbon Monoxide Gas Therapy.

As a promising therapeutic modality targeting cancer, gas therapy still faces critical challenges, especially in enhancing therapeutic efficacy and avoiding gas poisoning risks. Here, a pH/glutathione (GSH) dual stimuli-responsive CRISPR/Cas9 gene-editing nanoplatform combined with calcium-enhanced CO gas therapy for precise anticancer therapy, is established. In the tumor microenvironment (TME), the fast biodegradation of the CaCO3 layer via pH-induced hydrolyzation allows glucose oxidase (GOx) to catalyze glucose for H2 O2 production, which further reacts with manganese carbonyl (MnCO) and achieves the precise release of CO gas. Simultaneously, in situ Ca2+ overload from CaCO3 degradation disturbs mitochondrial Ca2+ homeostasis, resulting in Ca2+ -driven reactive oxygen species (ROS) formation and subsequent mitochondrial apoptosis signaling pathway activation. Subsequently, by GSH-induced cleavage of a disulfide bond, the released Cas9/sgRNA (RNP) can achieve nuclear factor E2-related factor 2 (Nrf2) gene ablation to sensitize gas therapy by interfering with ROS signaling. This therapeutic modality endows codelivery of CRISPR, ions, and gas with smart control features, which demonstrates great potential for future clinical applications in precise nanomedicine.

A new photodynamic therapy photosensitizer (p1) promotes apoptosis of keloid fibroblasts by targeting caspase-8

Photodynamic therapy (PDT) is a new therapy for treating cancer with less toxicity, high selectivity, good cooperativity, and repetitive usability. However, keloid treatment by PDT is mainly focused on clinical appearance, and few studies have been conducted on the mechanisms of PDT. In this study, key factors of the classical mitochondrial apoptosis signaling pathway were measured to assess the effect of a new PDT photosensitizer (p1). A specific inhibitor of caspase-8 (Z-IETD-FMK) was also used to verify the possible mechanisms. Twelve samples were obtained from 12 patients (six with keloids and six without) selected randomly from the Department of Plastic Surgery at Peking Union Medical College Hospital from January to December 2020. After cell culture, fibroblasts were divided into 13 groups. The morphology of fibroblasts in each group was observed by microscopy. Cell activity was measured by cell counting kit-8, and cell apoptotic morphology was observed by TUNEL staining. The reactive oxygen species (ROS) relative value was measured by a ROS test kit. The expression levels of key mitochondrial factors (caspase-3, caspase-8, cytochrome-c, Bax, and Bcl-2) were assessed by western blot, and mRNA expression of caspase-3 and caspase-8 was measured by RT-qPCR. We showed that p1 had a satisfactory proapoptotic effect on keloid fibroblasts by increasing the expression of ROS, caspase-3, caspase-8, and cytochrome-c, and decreasing the Bcl-2/Bax ratio; however, this effect was partially inhibited by Z-IETD-FMK, indicating that caspase-8 may be one of the p1’s targets to achieve the proapoptotic effect.

Role of endoplasmic reticulum stress in apoptosis induced by HK2 inhibitor and its potential as a new drug combination strategy.

Compared with normal cells, tumor cells mainly obtain energy through aerobic glycolysis. Hexokinase 2 (HK2) plays a key role in the regulation of tumor cell aerobic glycolysis, and targeting HK2 has become a new strategy for cancer treatment. However, little is known about the role of HK2 in colon cancer and the regulation of its targeted inhibitors. In this study, we found that the expression of HK2 in colorectal cancer tissues was significantly higher than that in adjacent tissues, and the expression level of HK2 in metastatic colorectal cancer was further increased. Meanwhile, the expression level of HK2 was closely related to clinical TNM stage and outcome of colorectal cancer patients. We provide here evidence that HK2 inhibitor 3-Bromopyruvate acid (3-BP) can significantly inhibit the survival and proliferation of colon cancer cells, and induce apoptosis through mitochondrial apoptosis signaling pathway. In addition, we found that 3-BP can also induce endoplasmic reticulum stress in colon cancer cells, the mechanism may be through the increase of intracellular calcium concentration. In vitro and in vivo experiments showed that inhibition of endoplasmic reticulum stress could further increase the proliferation inhibition and apoptosis induced by 3-BP. Collectively, our results show that HK2 is highly expressed in colorectal cancer. 3-BP, an inhibitor of HK2, can induce apoptosis and endoplasmic reticulum stress in colon cancer cells. Endoplasmic reticulum stress plays a protective role in cell death induced by 3-BP. This result suggested that targeting HK2 and endoplasmic reticulum stress may be a valuable strategy in targeted and combination therapy of colon cancer.

Dbdpe and Zno Nps Synergistically Induce Neurotoxicity of Sk-N-Sh Cells and Activate Mitochondrial Apoptosis Signaling Pathway and Nrf2-Mediated Antioxidant Pathway

Dbdpe and Zno Nps Synergistically Induce Neurotoxicity of Sk-N-Sh Cells and Activate Mitochondrial Apoptosis Signaling Pathway and Nrf2-Mediated Antioxidant Pathway

Pollution characteristics, exposure assessment and potential cardiotoxicities of PM2.5-bound benzotriazole and its derivatives in typical Chinese cities

Benzotriazole and its derivatives (BTRs), classified as high-volume production chemicals, have been widely detected in various environmental media, including the atmosphere, water, soil and dust, as well as organisms. However, studies on the pollution characteristics and health impact of PM2.5 related BTRs are so far limited. This study is the first to demonstrate the regional scale distribution of PM2.5-bound BTRs and their potential cardiotoxicities. Optimized methods of extraction, purification and GC-EI-MS/MS were applied to characterize and analyze PM2.5-bound BTRs from three cities in China during the winter of 2018. The concentration of ∑BTRs in Taiyuan (6.28 ng·m−3) was more than three times that in Shanghai (1.53 ng·m−3) and Guangzhou (1.99 ng·m−3). Benzotriazole (BTR) and 5-methyl-1H-benzotriazole (5TTR) contributed more than 80% of ∑BTRs concentration as the major pollutants among three cities. The correlation analysis indicated that there was a positive correlation between temperature and concentration of BTR and a negative correlation between temperature and concentration of 5TTR. In addition, the risk of BTRs exposure to toddlers should be paid more attention in Taiyuan by the human exposure assessment. Furthermore, toxicity screening by experimental methods indicated that 4-methyl-1H-benzotriazole (4TTR) was the most harmful to cardiomyocytes. The western blot assay showed a ROS-mediated mitochondrial apoptosis signaling pathway was activated after exposure to 4TTR in neonatal rat cardiomyocytes (NRCMs). On the other hand, metabolomics revealed that exposure of 4TTR to NRCMs disturbed mitochondrial energy metabolism by disturbing pantothenate and coenzyme A synthesis pathway. Our study not only clarifies the contamination profiles of PM2.5-bound BTRs in typical Chinese cities but also reveals their cardiotoxicities associated with mitochondrial dysfunction.

Sy-1365, a Potent and Selective CDK7 Inhibitor, Exhibits Anti-Tumor Activity in Preclinical Models of Hematologic Malignancies, and Demonstrates Interactions with the BCL-XL/BCL2 Mitochondrial Apoptosis Signaling Pathway in Leukemia

Introduction: The transcriptional kinase CDK7 has been implicated in the pathogenesis of multiple malignancies, including hematologic malignancies, and may play important roles in regulation of oncogenic transcriptional dependencies and in regulation of the mitochondrial apoptosis machinery in tumors. SY-1365, a selective inhibitor of CDK7, was developed to exploit tumor dependencies driven by CDK7 and is in clinical development in patients with advanced solid tumors. We have previously reported that SY-1365 selectively induces apoptosis in leukemic cells relative to non-malignant cells in vitro, and demonstrates anti-tumor activity in AML xenografts. Here we report that SY-1365 has potent inhibitory activity against a broad panel of hematologic malignancies in vitro and ex vivo, induces regressions in AML xenografts using a clinically relevant dosing regimen, and demonstrates synergy with the BCL2 inhibitor venetoclax in AML cells in vitro.Methods: SY-1365 dose-response curves were measured using the ATP-lite assay in a panel of 85 leukemia and lymphoma cell lines in vitro, and in a panel of 226 patient-derived tumor samples from 5 leukemic indications cultured ex vivo. In vivo tumor inhibitory activity was evaluated using once weekly (QW) and twice weekly (BIW) dosing regimens in ML-2 and Kasumi-1 AML xenografts. Growth-rate (GR) adjusted dose response curves were used to classify cell lines into low and high sensitivity groups, and were evaluated relative to RNA expression data to identify markers predictive of sensitivity to SY-1365. Synergy between SY-1365 and venetoclax was evaluated in vitro using KG-1 AML cells.Results: In 42 leukemia lines, SY-1365 had a median IC50 of 11nM (range 1-79nM), with 48% (20/42) demonstrating particularly low IC50s (<10nM). In 43 lymphoma lines, the median IC50 was 6nM (range 1-39nM), with 59% (27/46) demonstrating low IC50s (<10nM). Similarly, SY-1365 demonstrated potent inhibitory activity against patient-derived leukemia samples cultured ex vivo (median IC50 of 28nM), with 82% (185/226) of the samples showing IC50 below 100nM and 8% (19/226) under 10nM. Furthermore, analysis of growth rate (GR) in hematologic tumor cell lines showed that <10nM SY-1365 induced cytotoxicity (GR<0) in 19% (8/42) of leukemia and 21% (9/43) of lymphoma lines. To evaluate SY-1365 anti-tumor activity in vivo, 2 AML cell lines that demonstrate cytotoxic responses to SY-1365 in vitro (Kasumi-1 and ML-2) were grown as subcutaneous xenografts in mice. QW and BIW dosing of 40mg/kg SY-1365 induced tumor regressions in Kasumi-1 xenografts, and BIW dosing induced tumor stasis in ML-2 xenografts. Tumor growth inhibition was detected with a 20mg/kg BIW dosing regimen in both models. Comparison of SY-1365 sensitivity with RNA expression in hematologic tumor cell lines revealed that lower expression of BCL2L1, which encodes the mitochondrial apoptosis inhibitor BCL-XL, was highly predictive of sensitivity in leukemia cell lines (Accuracy=89%, FDR<0.05); no such relationship was observed in lymphoma lines. Considering the established role of BCL2 and the mitochondrial apoptosis pathway in leukemia, we reasoned that BCL-XL low-expressing leukemias, in addition to being sensitive to SY-1365, would also be reliant on BCL2 for survival. Indeed, in vitro combination studies demonstrated synergistic inhibitory activity of SY-1365 with the BCL2 inhibitor venetoclax in KG-1 AML cells. A potential synergistic interaction with SY-1365 and venetoclax is currently being explored in vivo with AML xenografts.Conclusions: SY-1365 shows potent in vitro inhibitory activity in multiple hematological indications and can induce AML xenograft tumor regression in mice. In vitro exploratory biomarker and tumor cell inhibitory combination studies in AML suggest a role for mitochondrial apoptosis signaling in mediating sensitivity to SY-1365 and support the evaluation of a combination with venetoclax. SY-1365 is currently being assessed in a phase 1 trial in adult patients with advanced solid tumors (NCT03134638). The results described here support the potential for additional exploration in patients with hematological malignancies. DisclosuresHodgson:Syros Pharmaceuticals: Employment, Equity Ownership. Johannessen:Syros Pharmaceuticals: Employment, Equity Ownership. Rajagopal:Syros Pharmaceuticals: Employment, Equity Ownership. Hu:Syros Pharmaceuticals: Employment, Equity Ownership. Orlando:Syros Pharmaceuticals: Employment, Equity Ownership. Eaton:Syros Pharmaceuticals: Employment, Equity Ownership. McKeown:Syros Pharmaceuticals: Employment. Tyner:Takeda Pharmaceutical Company: Research Funding; Agios Pharmaceuticals: Research Funding; Incyte Corporation: Research Funding; Gilead: Research Funding; Janssen Pharmaceutica: Research Funding; Syros: Research Funding; Aptose Biosciences: Research Funding; Seattle Genetics: Research Funding; Constellation Pharmaceuticals: Research Funding; Genentech: Research Funding; Array Biopharma: Research Funding; AstraZeneca: Research Funding; Leap Oncology: Consultancy. Sprott:Syros Pharmaceuticals: Employment, Equity Ownership. Fritz:Syros Pharmaceuticals: Employment, Equity Ownership. Di Tomaso:Syros Pharmaceuticals: Employment.

Tannic acid repair of zearalenone-induced damage by regulating the death receptor and mitochondrial apoptosis signaling pathway in mice

Zearalenone (ZEA) is an estrogenic toxin produced by Fusarium strains, that is widely present in crops, and endangers the reproductive system of animals. Tannic acid (TA) is a natural polyphenolic substance that is widespread in the roots, stems, and leaves of plants, and has special pharmacological activity. This study was designed to investigate the therapeutic effect of TA on ZEA-induced ovarian damage in mice and to explore the molecular mechanism involved. Ninety healthy Kunming female mice were divided into six equal groups. All the groups but the control group were administered daily with ZEA [10 mg/kg body weight (bw)] orally, for 7 days, to induce damage to the reproductive system. Some groups were also administered with TA (50, 100, and 200 mg/bw) for 7 days. Mice were euthanized 24 h later to allow for collection of serum and ovaries. TA can effectively alleviate the appearance of congestion and redness of the ovary, caused by ZEA, and increase the number of healthy growing follicles. Moreover, the estrogen content and the levels of MDA and ROS in the ovaries can be effectively reduced by TA. It can also reduce the apoptosis of ovarian cells, decreases the protein expression of the estrogen receptor, Fas, Fasl, caspase-3, caspase-8, caspase-9, and Bax, and increases the protein expression of Bcl-2. Our study indicates that TA reduces the strong estrogen and oxidative damage induced by ZEA, and these therapeutic effects may be partially mediated by the death receptor and mitochondrial apoptosis signaling pathway.

Copper induces mitochondria-mediated apoptosis via AMPK-mTOR pathway in hypothalamus of Pigs

Copper (Cu), one of the heavy metals, is far beyond the carrying capacity of the environment with Cu mining, industrial wastewater discharging and the use of Cu-containing pesticides. Intaking excess Cu can cause toxic effects on liver, kidney, heart, but few studies report Cu toxicity on brain tissue. It is noteworthy that most toxicity tests are based on rodent models, but large mammals chosen as animal models has no reported. To explore the relationship of the Cu toxicity and mitochondria-mediated apoptosis on hypothalamus in pigs, the content of Cu, histomorphology, mitochondrial related indicators, apoptosis, and AMPK-mTOR signaling pathway were detected. Results showed that Cu could accumulate in hypothalamus and lead to mitochondrial dysfunction, evidenced by the decrease of ATP production, activities of respiratory chain complex I-IV, and mitochondrial respiratory function in Cu-treated groups. Additionally, the genes and proteins expression of Bax, Caspase-3, Cytc in treatment group were higher than control group. Furthermore, the protein level of p-AMPK was enhanced significantly and p-mTOR was declined, which manifested that AMPK-mTOR signaling pathway was activated in Cu-treated groups. In conclusion, this study illuminated that the accumulation of Cu could cause mitochondrial dysfunction, induce mitochondria-mediated apoptosis and activate AMPK-mTOR pathway in hypothalamus.

Gegen Qinlian Decoction Ameliorates Hyperuricemia-Induced Renal Tubular Injury via Blocking the Inflammatory Signaling Pathway.

Background: Gegen Qinlian decoction (GGQLD) is a typical traditional Chinese medicine (TCM) prescription documented in Shang Han Lun. Clinically, GGQLD has been utilized to manage the inflammatory symptoms of metabolic diseases and to protect against renal damage in China. In the present study, a hypothesis was proposed that the multi-target solution of GGQLD produced anti-inflammatory effects on ameliorating hyperuricemia (HUA). Methods: A total of 30 primary HUA patients receiving GGQLD treatment (two doses daily) for 4 weeks were selected. Then, differences in uric acid (UA) levels and expression of peripheral blood mononuclear cells (PBMCs) and urinary exosomes before and after treatment were analyzed. The therapeutic indexes for the active ingredients in GGQLD against HUA were confirmed through pharmacological subnetwork analysis. Besides, the HUA rat model was established through oral gavage of potassium oxonate and treated with oral GGQLD. In addition, proximal tubular epithelial cells (PTECs) were stimulated by UA and intervened with GGQLD for 48 h. Subsequently, RNA-seq, flow cytometry, and confocal immunofluorescence microscopy were further conducted to characterize the differences in UA-mediated inflammation and apoptosis of human renal tubular epithelial cells pre- and post-administration of GGQLD. In the meanwhile, quantitative real-time PCR (qPCR) was carried out to determine gene expression, whereas a western blotting (WB) assay was conducted to measure protein expression. Results: Our network analysis revealed that GGQLD treated HUA via the anti-inflammatory and antiapoptotic pathways. Additionally, NLPR3 expression significantly decreased in PBMCs and urinary exosomes of HUA patients after GGQLD treatment. In vivo, GGQLD treatment alleviated HUA-induced renal inflammation, which was associated with decreased expression of NLRP3 inflammasomes and apoptosis-related mRNAs. Moreover, GGQLD promoted renal UA excretion by inhibiting the activation of GSDMD-dependent pyroptosis induced by NLRP3 inflammasomes and by reducing apoptosis via the mitochondrial apoptosis signaling pathway in vitro. Conclusion: This study indicates that GGQLD efficiently reduces inflammatory responses while promoting UA excretion in HUA. Our findings also provide compelling evidence supporting the idea that GGQLD protects against the UA-mediated renal tubular epithelial cell inflammation through the mitochondrial apoptosis signaling pathways. Taken together, these findings have demonstrated a novel therapeutic method for the treatment of HUA.

The Flavonol Isoquercitrin Promotes Mitochondrial-Dependent Apoptosis in SK-Mel-2 Melanoma Cell via the PI3K/AKT/mTOR Pathway.

Isoquercitrin (IQ), a major flavonol present in Prunus mume fruit, has gained much attention in recent studies because of its superior bioavailability and physiological effects. In this study, the anti-cancer mechanism of IQ against human melanoma, particularly its effect on the mitochondria-mediated apoptosis, was investigated. Treatment with IQ at 25 μM concentration effectively inhibited the proliferation of SK-MEL-2 skin cancer cells while the same concentration did not exhibit cytotoxicity against human keratinocytes HaCaT. Morphological analysis and clonogenic assay also showed that IQ can alter the growth and long-term survival of SK-MEL-2 cells. IQ also induced apoptosis in the melanoma cells as manifested in the nuclear morphology changes, DNA fragmentation, increase in the apoptosis rate (17.69% at 25 μM) and accumulation of sub-G1 cell cycle phase population (19.55% at 25 μM). Western blot analysis revealed the involvement of the mitochondrial apoptosis signaling pathway in the anti-cancer property of IQ. Treatment with IQ resulted in the decrease in the levels of procaspase-8 and -9, and Bcl-2 protein, and an increase in the expression of cleaved PARP and Bax. Moreover, AIF and Endo G protein expression increased, indicating a caspase-independent mitochondrial-mediated apoptosis. The anti-proliferative activity of IQ against SK-MEL-2 can also be attributed to the downregulation of the PI3K/AktmTOR signaling pathway. These findings showed that IQ can be developed into a chemopreventive therapeutic agent against the melanoma cells.

Synergetic inhibition of daidzein and regular exercise on breast cancer in bearing-4T1 mice by regulating NK cells and apoptosis pathway

The aim of this study was to investigate the inhibition of daidzein or/and regular exercise on breast cancer and to reveal the potential biological mechanisms. BALB/c mice pretreated with regular exercise training for 20 days (15 m/min, 60 min/d) were orthotopically transplanted with mouse breast cancer cells (4T1), and then treated with daidzein (145 mg/kg) by gavage for another 22 days. Results showed that exercise or daidzein inhibited tumor growth in mice to a different degree. Particularly, co-treatment with exercise and daidzein showed an obviously synergistic inhibition on the tumor growth (P < 0.01), compared with the tumor control. Further researches indicated that the combination of exercise and daidzein synergistically mobilized and redistributed natural killer cells through upregulating the level of epinephrine and interleukin-6. Moreover, exercise combined with daidzein induces apoptosis in cancer cells via Fas/FasL-initiated mitochondrial apoptosis signaling pathway. These results suggested that regular exercise combined with daidzein may explore a candidate way to prevent and treat the breast cancer.