scholarly journals Protective Effects of Astragalus Polysaccharide on Sepsis-Induced Acute Kidney Injury

2021 ◽  
Vol 2021 ◽  
pp. 1-13
Author(s):  
Jie Sun ◽  
Shanzhai Wei ◽  
Yilai Zhang ◽  
Jia Li
Keyword(s):  
Caspase 3 ◽  
Morphological Changes ◽  
Kidney Injury ◽  
Cell Counting ◽  
Enzyme Linked Immunosorbent Assay ◽  
In Vitro Experiments ◽  
Caspase 9 ◽  
Astragalus Polysaccharide ◽  
Tnf Α

Objective. To explore the protective roles of Astragalus polysaccharide (APS) on acute renal injury (AKI) induced by sepsis. Methods. Firstly, an animal model of sepsis-induced AKI was established by injecting lipopolysaccharide (LPS) into mice. The mice were pretreated with an intraperitoneal injection of 1, 3, and 5 mg/(kg·d) APS for 3 consecutive days. The severity of kidney injury was then scored by histopathological analysis, and the concentrations of serum urea nitrogen (BUN) and serum creatinine (SCr) and the levels of tumor necrosis factor α (TNF-α) and interleukin-1β (IL-1β) were determined as well. In in vitro experiments, lipopolysaccharide (LPS) was used to induce HK-2 cell injury to establish a sepsis-induced AKI cell model, and the cell counting kit-8 (CCK-8) method was performed to determine the cytotoxicity and appropriate experimental concentration of APS. Then, cells were divided into the control, LPS, and APS+LPS groups. Cell apoptosis and inflammation-related TNF-α, IL-1β, IL-6, and IL-8 were determined by flow cytometry and enzyme-linked immunosorbent assay (ELISA), respectively. The microscope was used to observe the morphological changes of cells, and the cell migration ability was measured by wound healing assay. RT-qPCR and Western blot assay were used to determine the mRNA and protein levels of apoptosis-related factors including caspase-3, caspase-9, Bax, and Bcl-2; endoplasmic reticulum stress- (ERS-) related biomarkers including C/EBP homologous protein (CHOP) and glucose-regulated protein78 (GRP78); and epithelial-mesenchymal transition- (EMT-) related biomarkers including E-cadherin, Snail, α-smooth muscle actin (α-SMΑ), and Vimentin. Results. In vivo experiments in mice showed that APS can reverse LPS-induced kidney damage in a concentration-dependent manner ( P < 0.05 ); the concentrations of BUN and Scr were increased (all P < 0.05 ); similarly, the levels of TNF-α and IL-1β were increased as well (all P < 0.05 ). In in vitro experiments, the results showed that LPS can significantly cause HK-2 cell damage and induce apoptosis, inflammation, ERS, and EMT. When APS concentration was in the range of 0-200 μg/mL, it had no cytotoxicity in HK-2 cells, and 100 μg/mL APS pretreatment could significantly mitigate the decrease of cell activity induced by LPS ( P < 0.05 ). Compared with the LPS group, APS pretreatment could inhibit the expression of inflammatory factors including TNF-α, IL-1 β, IL-6, and IL-8 (all P < 0.05 ), reducing the number of apoptotic cells ( P < 0.05 ), suppressing the expression of caspase-3, caspase-9, and Bax, but upregulating the expression levels of Bcl-2. In ERS, APS pretreatment inhibited LPS-induced upregulation of CHOP and GRP78. Moreover, in EMT, APS pretreatment could inhibit the morphological changes of cells, downregulate the migration, decrease the expression of EMT biomarkers, and inhibit the process of EMT. Conclusion. APS could alleviate sepsis-induced AKI by regulating inflammation, apoptosis, ERS, and EMT.

scholarly journals Ketamine induces endoplasmic reticulum stress in rats and SV-HUC-1 human uroepithelial cells by activating NLRP3/TXNIP aix

2019 ◽  
Vol 39 (10) ◽  
Author(s):  
Lingjuan Cui ◽  
Xiaoyan Jiang ◽  
Chengjun Zhang ◽  
Danxia Li ◽  
Shengqiang Yu ◽  
...  
Keyword(s):  
Caspase 3 ◽  
B Cell Lymphoma ◽  
Cell Counting ◽  
Enzyme Linked Immunosorbent Assay ◽  
Interacting Protein ◽  
Protein Levels ◽  
Tnf Α ◽  
Uroepithelial Cells

Abstract Many clinical studies have been conducted on ketamine-associated cystitis. However, the underlying mechanisms of ketamine-associated cystitis still remain unclear. Bladder tissues of rats were stained by Hematoxylin and Eosin (HE). The viability of human uroepithelial cells (SV-HUC-1 cells) was determined by cell counting kit-8 (CCK-8). Apoptosis and reactive oxygen species (ROS) were examined by flow cytometry. Additionally, the expressions of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), IL-1β and IL-18 were respectively determined by reverse transcription quantitative (RTq)-PCR and enzyme-linked immunosorbent assay (ELISA). The mRNA and protein levels of B-cell lymphoma/leukemia-2 (Bcl2), Bcl-2-associated X protein (Bax), cleaved caspase 3, glucose-regulated protein 78 (GRP78), CCAAT/enhancer binding protein homologous protein (CHOP), NOD-like receptor 3 (NLRP3), thioredoxin-interacting protein (TXNIP), Catalase and MnSOD were examined by RT-qPCR and Western blot. Small interfering RNA target TXNIP transfection was performed using Lipofectamine™ 2000. We found that ketamine effectively damaged bladder tissues of rats and promoted apoptosis through regulating the expression levels of GRP78, CHOP, Bcl-2, Bax and cleaved Caspase-3 proteins in vivo and in vitro. NLRP3 inflammatory body and TXNIP were activated by ketamine, which was supported by the changes in TNF-α, IL-6, IL-1 and IL-18 in vivo and in vitro. Furthermore, knocking down TXNIP reversed the effects of ketamine on apoptosis and NLRP3 inflammatory body in SV-HUC-1 cells. Meanwhile, the changes of Catalase and MnSOD showed that ROS was enhanced by ketamine, however, such an effect was ameliorated by down-regulation of TXNIP in SV-HUC-1 cells. Ketamine promoted cell apoptosis and induced inflammation in vivo and in vitro by regulating NLRP3/TXNIP aix.

scholarly journals Direct Effect of Septic Plasma in Human Cell Lines Viability

2018 ◽  
Vol 47 (1-3) ◽  
pp. 270-276
Author(s):  
Grazia Maria Virzì ◽  
Chiara Borga ◽  
Chiara Pasqualin ◽  
Silvia Pastori ◽  
Alessandra Brocca ◽  
...  
Keyword(s):  
Cytochrome C ◽  
Cell Viability ◽  
Cell Lines ◽  
Caspase 3 ◽  
Test Group ◽  
Organ Injury ◽  
Enzyme Linked Immunosorbent Assay ◽  
Caspase 9 ◽  
Renal Tubular Cells

Background: Sepsis is a life-threatening condition often associated with a high incidence of multiple organs injury. Several papers suggested the immune response by itself, with the production of humoral inflammatory mediators, is crucial in determining organ injury. However, little is known of how sepsis directly induces organ injury at the cellular levels. To assess this point, we set up an in vitro study to investigate the response of renal tubular cells (RTCs), monocytes (U937) and hepatocytes (HepG2) after 24 h-incubation with septic patients’ plasma. Methods: We enrolled 26 septic patients (“test” group). We evaluated cell viability, apoptosis and necrosis by flow cytometer. Caspase-3,-8,-9 and cytochrome-c concentrations have been analyzed using the Human enzyme-linked immunosorbent assay kit. Results: We found that a decrease of cell viability in all cell lines tested was associated to the increase of apoptosis in RTCs and U937 (p < 0.0001) and increase of necrosis in HepG2 (p < 0.5). The increase of apoptosis in RTCs and U937 cells was confirmed by higher levels of caspase-3 (p < 0.0001). We showed that apoptosis in both RTCs and U937 was triggered by the activation of the intrinsic pathway, as caspase-9 and cytochrome-c levels significantly increased (p < 0.0001), while caspase-8 did not change. This assumption was strengthened by the significant correlation of caspase-9 with both cytochrome-c (r = 0.73 for RTCs and r = 0.69 for U937) and caspase-3 (r = 0.69 for RTCs and r = 0.63 for U937). Conclusion: Humoral mediators in septic patients’ plasma induce apoptosis. This fact suggests that apoptosis inhibitors should be investigated as future strategy to reduce sepsis-induced organ damages.

scholarly journals Shenshuaikang Enema, a Chinese Herbal Remedy, Inhibited Hypoxia and Reoxygenation-Induced Apoptosis in Renal Tubular Epithelial Cells by Inhibiting Oxidative Damage-Dependent JNK/Caspase-3 Signaling Pathways Using Network Pharmacology

2020 ◽  
Vol 2020 ◽  
pp. 1-12
Author(s):  
Hongmei Lu ◽  
Xinyi Luo ◽  
Yuhua He ◽  
Bo Qu ◽  
Liangbin Zhao ◽  
...  
Keyword(s):  
Oxidative Damage ◽  
Caspase 3 ◽  
Cell Counting ◽  
Network Pharmacology ◽  
In Vitro Experiments ◽  
Chinese Herbal ◽  
Cell Vitality ◽  
Target Network ◽  
Renal Tubular

Background. Acute kidney injury (AKI) is a common clinically critical illness with serious consequences for the patients. Shenshuaikang enema (SE) is a Chinese herbal compound that is used to treat AKI in clinical practice. However, its mechanism of action remains unclear. Aim. The aim of this study was to investigate the therapeutic effect of SE and explore the molecular mechanisms using network pharmacology and in vitro experiments. Materials and Methods. The herb-component-target network was constructed based on network pharmacology. The predicted targets and pathways were validated using in vitro experiments. A renal tubular epithelial cell line (HK-2 cells) was exposed to hypoxia and reoxygenation (H/R) using air-tight conditions for five hours and treated with different concentrations of SE (25%, 50%, and 75%) to assess cell viability and apoptosis and determine the optimal experimental dose. Subsequently, H/R-injured HK-2 cells were pretreated with the optimal SE dose and then randomly divided into three groups, the SE, SE-SP600125 (inhibitor of JNK), and SE-NAC (antioxidant) groups. The cell vitality, apoptosis, and death were evaluated using the cell counting kit 8 (CCK8) and carboxyfluorescein succinimidyl ester/propidium iodide (CFSF/PI) staining. The apoptosis-related protein JNK and Caspase-3 were assessed by Western blot. Expression of JNK and Caspase-3 genes was analyzed using real-time quantitative polymerase chain reaction (RT-qPCR). Results. 123 active components and 226 targets were identified from four herbs that composed the herb-compound-target network based on transcriptomics and network pharmacology analyses. The KEGG pathway analyses revealed that the mitochondrial apoptosis pathway was involved in the therapeutic AKI effects of SE. Cell vitality of H/R-induced HK-2 cells was obviously increased when treating them with SE, and the apoptosis was significantly inhibited, especially in the SE (50%) group at 4 and 12 h after modeling. Pretreatment with antioxidant NAC obviously prevented cell death compared to the SE (50%) group, while no obvious reduction of apoptosis was observed in the SP600125 group. JNK expression level was significantly increased in the SE (50%) group compared to the SP600125 ( P < 0.01 ) and the NAC group ( P < 0.05 ). Caspase-3 was downregulated in the SE (50%) group compared to the SP600125 ( P < 0.01 ) and NAC group ( P < 0.05 ). Caspase-3 activation in the SP600125 group was higher than that in the NAC group ( P < 0.05 ). Moreover, the oxidative damage-dependent JNK/Caspase-3 pathway was identified in the H/R-injured HK-2 cells by inhibiting the JNK activation and oxidative damage. Conclusions. Our findings suggested that the H/R-triggered apoptosis in HK-2 cells was abrogated by SE by upregulating the oxidative damage-dependent JNK to trigger suppression of Caspase-3.

P0528RENOPROTECTIVE EFFECT OF IL-34 INHIBITION ON CISPLATIN-INDUCED NEPHROTOXICITY IN MICE

2020 ◽  
Vol 35 (Supplement_3) ◽  
Author(s):  
Yukihiro Wada ◽  
Masayuki Iyoda ◽  
Kei Matsumoto ◽  
Taihei Suzuki ◽  
Ken Iseri ◽  
...  
Keyword(s):  
Caspase 3 ◽  
Kidney Injury ◽  
Neutralizing Antibody ◽  
Inhibitory Effects ◽  
Tubulointerstitial Injury ◽  
Cisplatin Nephrotoxicity ◽  
Physiological Properties ◽  
Tnf Α ◽  
Vehicle Treatment

Abstract Background and Aims Interleukin (IL)-34, a macrophage (Mø) mediator, is expressed by tubular epithelial cells (TECs). However, the influence of IL-34 on TECs injury has not been fully elucidated. We investigated the physiological properties of IL-34 on TECs damage caused by cisplatin-nephrotoxicity (CP-N). Method 7-week-old male C57BL/6 (B6) mice (n=16) were fasted for 8 hours and then induced CP-N by intraperitoneal injection (IP) of CP (25 mg/kg) on day 0. Groups of animals were given either anti-mouse IL-34 antibody (CP+anti-IL-34 Ab, 400 ng/kg, n=8) or vehicle (CP+V, equal volume of saline, n=8) daily by IP from day -1 to day 2. Three age-matched male B6 mice were used as normal control (NC). All mice were sacrificed on day 3. In addition, mouse renal proximal TECs (MRTEpiC) were cultured to analyze the inhibitory effects of IL-34 on CP-induced TEC apoptosis. Cells were stimulated with CP (2 μg/mL), then treated with or without anti-IL-34 Ab (1000 pg/mL). Results Compared to the NC, CP+V mice exhibited marked acute kidney injury (AKI) and upregulated expression of IL-34 and its two receptors, cFMS and PTP-ζ. Compared to the vehicle treatment, anti-IL-34 Ab treatment significantly suppressed the intrarenal expression levels of IL-34 and its two receptors in CP-N mice; it also significantly suppressed serum IL-34 levels (72.1 ± 5.6 vs. 40.4 ± 7.5 pg/mL, p=0.013). Additionally, treatment with anti-IL-34 Ab significantly improved serum Cr levels (1.3 ± 0.2 vs. 0.7 ± 0.1 mg/mL, p=0.033), ameliorated tubulointerstitial injury (numbers of casts/HPF: 11.9 ± 2.6 vs. 6.5 ± 1.8, p=0.048), and suppressed the number of F4/80+ Mø (17.5 ± 2.7 vs. 11.1 ± 1.1/HPF, p=0.041) and TUNEL+ apoptotic cells (29.2 ± 4.9 vs. 16.7 ± 2.7/HPF, p=0.036) in CP-N mice. The renal cortical transcript levels of Kim-1, MIP-1/CCL3, TNF-α, and Bax were significantly lower in the CP+anti-IL-34 Ab mice than in the CP+V mice. Furthermore, the CP+anti-IL-34 Ab mice showed significantly less renal infiltration of CD11b+F4/80+TNF-α+ cells. In vitro, stimulation with CP induced the expression of IL-34 and its two receptors in MRTEpiC. Treatment with anti-IL-34 Ab significantly suppressed CP-induced caspase-3 and Bax expression with degradation of ERK1/2 phosphorylation in the damaged MRTEpiC. Conclusion IL-34 secreted from damaged TECs was involved in the progression of CP-N. Inhibition of IL-34 with neutralizing antibody directly prevented CP-induced TEC apoptosis by inhibiting the phosphorylation of ERK1/2. Blocking of IL-34 might suppressed proliferation of cytotoxic Mø, which indirectly led to the attenuation of CP-N. Thus, IL-34 represents a potential as therapeutic target for AKI with TECs injury.

scholarly journals Guaiane-Type Sesquiterpenoids From Dendranthema morifolium (Ramat.) S. Kitam Flowers Protect H9c2 Cardiomyocyte From LPS-Induced Injury

2019 ◽  
Vol 14 (7) ◽  
pp. 1934578X1986417
Author(s):  
Beibei Zhang ◽  
Mengnan Zeng ◽  
Meng Li ◽  
Wenjing Chen ◽  
Benke Li ◽  
...  
Keyword(s):  
Caspase 3 ◽  
Creatine Phosphate ◽  
Cardiomyocyte Apoptosis ◽  
Exocyclic Double Bond ◽  
Enzyme Linked Immunosorbent Assay ◽  
Protective Effects ◽  
Caspase 9 ◽  
Necrosis Factor Alpha ◽  
Thiazolyl Blue Tetrazolium Bromide ◽  
Tnf Α

This study investigated the protective effects of guaiane-type sesquiterpenoids isolated from Dendranthema morifolium (Ramat.) S. Kitam flowers on lipopolysaccharide (LPS)-induced injury in H9c2 cardiomyocyte. Cell viability was determined by thiazolyl blue tetrazolium bromide (MTT). The content of released tumor necrosis factor alpha (TNF- α) and interleukin 6 (IL-6) was evaluated by enzyme-linked immunosorbent assay. The levels of lactate dehydrogenase (LDH) and creatine phosphate kinase (CK) were measured by using commercial available kits. The protein expression levels of pelF2 α, GRP78, Bax, caspase-3, caspase-9, Bcl-2, LC3-II, and p62 were measured by in-cell Western. Flow cytometry was used to detect H9c2 cardiomyocyte apoptosis. Compounds 5, 7, 1, 8, and 2 exhibited the effects of cardioprotection and activity sequence enhancement. The levels of IL-6, TNF- α, LDH, CK, pelF2 α, GRP78, Bax, caspase-3, caspase-9, p62, and H9c2 cardiomyocyte apoptosis were increased in LPS-treated H9c2 cardiomyocyte, while those of Bcl-2 and LC3-II were decreased. These effects could be effectively reversed by compounds 5, 7, 1, 8, and 2. Results demonstrated that the guaiane-type sesquiterpenoids could prevent LPS-induced injury in cardiomyocyte by decreasing endoplasmic reticulum (ER) stress, apoptosis, and autophagy as well as downregulating the inflammatory mediators. In addition, the active groups of guaiane-type sesquiterpenoids might be the angelate at C-8 and the exocyclic double bond at C-11.

P0538HYPERIN PREVENTS ISCHEMIC/REPERFUSION AKI VIA STK40

2020 ◽  
Vol 35 (Supplement_3) ◽  
Author(s):  
Yan Xu ◽  
Yue Zhang
Keyword(s):  
Caspase 3 ◽  
Ischemia Reperfusion ◽  
Normal Group ◽  
Sham Operation ◽  
In Vitro Experiments ◽  
Rt Pcr ◽  
Sham Operation Group ◽  
Tnf Α ◽  
Operation Group

Abstract Background and Aims Ischemia-reperfusion injury (IRI) is the outcome of an inflammatory process and tubular cell death that is triggered by undergoing a transient reduction or cessation of blood flow and following by reperfusion. Unresolved IRI can contribute to chronic kidney disease even death. Our aims is to investigate the protective effect of hyperin on ischemia-reperfusion renal injury (IRI) and its possible mechanism. Method ① The transcriptome chip data of multiple IRI models were selected from the NCBI GEO DateSets database and a number of key proteins that could participate in IRI were screened out (the fold increase was greater than 2 fold and was statistically significant). Network and transcript binding motif analysis was performed to determine the best binding protein. ② C57BL / 6J mice were selected and randomly divided into normal group, sham operation group, IRI group (bilateral renal pedicle clamping for 45min), hyperin + IRI group (50mg / kg.d per day, 7 days before surgery ), DMSO + IRI group (7 days before the operation, the same amount of DMSO was administered to the stomach every day, and the operation was the same as AKI), with 6 rats in each group. Renal tissue and blood were collected 24 hours after operation for testing. ③ In vitro experiments, human proximal tubule epithelial cells (HK-2) were selected and divided into hypoxia 3, 6, 9, 12, 24, 36, and 48h for reoxygenation of 1, 3, and 6h respectively. Relevant indicators for RT-PCR detection were determined Optimal hypoxia time. The drug safe concentration was selected according to 0, 5, 10, 25, 50, 100, 200, 400 μg / ml hyperin pre-treatment for 12 hours, and the CCK8 reagent was added for 2 hours to measure the absorbance at 450 nm. The cells were randomly divided into normal group, hypoxia group, hypoxia + DMSO group, hypoxia + hyperin group, and related indexes were detected by RT-PCR and Western Blot. ④ Obtain the tertiary structure of the protein and the three-dimensional structure of the hyperin molecule from the RCSB Protein Data Bank website and the PubChem compound database, and use molecular docking technology to determine the proteins that can bind to hyperin using autodock software and analyze their binding ability. Results Bioinformatics analysis suggested that STK40 protein is one of the key factors of IRI and may be a target for preventing and treating diseases. In vivo experiments showed that compared with the normal group and the sham operation group, the levels of serum creatinine, blood urea nitrogen, and kim-1 in rats were significantly increased after AKI, and HE staining of pathological sections showed an increase in renal tubular injury scores. Significantly decreased (P&lt;0.05); RT-PCR results showed that kim-1, caspase-3, NF-κB, IL-6, TNF-α increased significantly after AKI, STK40, Bcl2 / BAX decreased, and the above after hyperin The indicators changed in opposite directions (P &lt;0.05). In vitro experiments: The best time for hypoxia is 24h hypoxia + 1h reoxygenation; compared with the control group, the drug concentration is &lt;100 μg / mL and the cell proliferation activity rate is&gt; 90%, so the hyperin concentration was selected as 50 μg / mL (P &lt; 0.05); RT-PCR results showed that Hif1-α, caspase-3, NF-κB, IL-6, TNF-α significantly increased, and STK40, Bcl2 / BAX decreased compared with the normal group. After administration of hyperin, the above indexes changed in opposite directions (P &lt;0.05). Conclusion In this study, using molecular docking technology and constructing IRI mice model, it was confirmed that hyperin can reduce IRI and exert a protective effect on IRI by inhibiting STK40 expression.

scholarly journals Meisoindigo inhibits cellular proliferation via down-regulation of the PI3K/Akt pathway and induces cellular apoptosis in glioblastoma U87 cells

2021 ◽  
Author(s):  
Lijuan Gu ◽  
Yi Zhou ◽  
Yingze Ye ◽  
Xiqun Zhu ◽  
Tong Jin ◽  
...  
Keyword(s):  
Caspase 3 ◽  
Cellular Proliferation ◽  
Nuclear Translocation ◽  
Morphological Changes ◽  
Inhibitory Effect ◽  
Cell Counting ◽  
Akt Pathway ◽  
Dependent Manner ◽  
Caspase 9 ◽  
U87 Cells

Objective: The current study was to explore whether meisoindigo was effective in suppressing proliferation and inducing apoptosis of human glioblastoma multiforme U87 cells and to explore its possible mechanisms. Method: Morphological changes were observed by light microscopy. Cell counting kit-8 (CCK-8) assay was performed to detect cellular proliferation. Apoptosis was monitored by flow cytometry. Akt, phospho-Akt, PI3K, p65, phospho-p65 and apoptosis-related proteins caspase-3 and caspase-9 were examined by Western blotting assays. Immunofluorescence was used to evaluate level of P65 expression in cells. Result: Meisoindigo inhibited the proliferation of U87 cells, and the inhibitory effect increased in a dose dependent manner. Moreover, meisoindigo exposure triggered an increase in the level of caspase-3 and caspase-9, supporting its role in the activation of apoptosis. Furthermore, meisoindigo reduced the expression of PI3K, Akt, phospho-Akt, NF-κB, p65 and phospho-p65 in U87 cells, and displacement of p65 from the nucleus to the cytoplasm. Conclusion: Meisoindigo inhibits proliferation and induces apoptosis of U87 cells, probably through down-regulating the PI3K/Akt pathway and reducing nuclear translocation of NF-κB p65.

Protective effect of dihydromyricetin on vascular smooth muscle cell apoptosis induced by hydrogen peroxide in rats

Perfusion ◽  
2022 ◽  
pp. 026765912110599
Author(s):  
Wenling Li ◽  
Hua Sang ◽  
Xin Xu ◽  
Yuanyuan Zhang ◽  
Xiangying Meng ◽  
...  
Keyword(s):  
Hydrogen Peroxide ◽  
Smooth Muscle ◽  
Vascular Smooth Muscle ◽  
Protective Effect ◽  
Cell Apoptosis ◽  
Caspase 3 ◽  
Morphological Changes ◽  
Ros Scavenging ◽  
Caspase 9

Objective Dihydromyricetin (DMY), also called Ampelopsin, which was extracted from Ampelopsis grossedentata, has been demonstrated to have a protective effect against cell oxidative injury and cell apoptosis in vitro. In the present study, we tried to study the role of DMY on apoptosis of vascular smooth muscle cells (VSMCs) induced by hydrogen peroxide (H2O2) and explore the underlying mechanisms. Methods Apoptotic cells were detected by Hematoxylin and Eosin (H.E.) staining, Hoechst 33342 staining, and Annexin V-fluorescein isothiocyanate binding assay. The intracellular reactive oxygen species (ROS) level was estimated through fluorescence assay. The mRNA and protein expression of Caspase-3, Caspase-9, Bcl-2, and Bax were determined by reverse transcription-polymerase chain reaction (RT-PCR) and western blot. Results The results showed that the pretreatment of VSMCs with DMY not only significantly increased cell viability, reduced intracellular ROS release, alleviated the morphological changes of apoptosis, and decreased the apoptosis rate, but also upregulated Bcl-2 expression and downregulated Caspase-3, Caspase-9, Bax expression, and ultimately attenuated the H2O2-stimulated apoptosis. Conclusion The inhibition of DMY on VSMC apoptosis may be mediated by ROS scavenging and the activation of the mitochondrial apoptotic signaling pathway.

Role of midkine in cadmium-induced liver, heart and kidney damage

2010 ◽  
Vol 30 (5) ◽  
pp. 391-397 ◽  
Author(s):  
Nuray Yazihan ◽  
Mehtap Kacar Kocak ◽  
Ethem Akcil ◽  
Onur Erdem ◽  
Ahmet Sayal
Keyword(s):  
Messenger Rna ◽  
Caspase 3 ◽  
Biological Effects ◽  
Toxic Substance ◽  
Underlying Mechanism ◽  
Enzyme Linked Immunosorbent Assay ◽  
Activity Levels ◽  
Cd Toxicity ◽  
Tnf Α

Accumulation of the widespread environmental toxin cadmium (Cd) in tissues results in toxicity. Cd, which can induce a broad spectrum of biological effects, is a toxic substance and is associated with inflammation and apoptosis. Midkine (MK) has fibrinolytic, antiapoptotic, transforming, angiogenetic and chemotactic activities. After Cd toxicity, we found increased MK expression in liver cells in an in vitro cell culture model. The aim of this study was to determine the possibility of relationship between tissue MK expression levels, tumor necrosis factor α(TNF-α) levels and apoptosis in a chronic Cd toxicity model in rats. Male Wistar rats were exposed to Cd at the dose of 15 parts per million (ppm) for 8 weeks. MK levels were measured in kidney, heart and liver tissue by enzyme-linked-immunosorbent assay (ELISA). MK messenger RNA (mRNA) expression was evaluated by RT-PCR. Tissue apoptosis level was evaluated with tissue caspase-3 activity levels. Accumulation of Cd in liver is higher than the kidney and heart. Cd-treated rats had significantly higher tissue TNF-α and caspase-3 levels when compared with the control rats (p < 0.001). MK mRNA and protein levels were also significantly upregulated in the Cd-treated group (p < 0.05, p < 0.001, respectively). When compared with apoptosis in tissues, it was more prominent in the liver than kidney and heart. MK level is found increased 3, 1.7 and 1.3× folds in liver, kidney and heart, respectively. Our results showed that chronic Cd administration induces inflammation and apoptosis in rat liver, kidney and heart. MK involved in damage mechanisms of Cd-induced tissues. Further studies will show the underlying mechanism of increased MK expression in Cd toxicity.

scholarly journals Wogonoside exerts potential anti-tumor activity against bladder cancer in vivo and in vitro via regulation of GSK3β/ERK/AKT signaling pathway

2021 ◽  
Vol 18 (7) ◽  
pp. 1385-1390
Author(s):  
Jiexiang Chen ◽  
Yong Cao ◽  
Yan Li ◽  
Li Tang ◽  
Xiaolan Yu ◽  
...  
Keyword(s):  
Bladder Cancer ◽  
Antitumor Activity ◽  
Cell Line ◽  
Proliferative Activity ◽  
Caspase 3 ◽  
B Cell Lymphoma ◽  
Enzyme Linked Immunosorbent Assay ◽  
Caspase 9 ◽  
Gsk 3Β

Purpose: To explore the antitumor activity of wogonoside on bladder cancer, and its underlying mechanism of action. Methods: Methyl thiazolyl tetrazolium (MTT) assay was applied to determine the anti-proliferative activity of wogonoside (2 - 128 μM) on bladder cancer 5637 cell line at various times, and the halfmaximal inhibitory concentration (IC50) was measured. The antitumor activity of wogonoside (30 mg/kg, ip) against bladder cancer 5637 cell line was evaluated in nude mice bearing human bladder cancer 5637 cells. Additionally, western blotting and enzyme-linked immunosorbent assay (ELISA) were carried out to investigate the levels of the caspase-3, caspase-9, B cell lymphoma/leukemia-2 (Bcl-2), Bcl-2 associated X-protein (Bax), phosphorylated (p)-glycogen synthase kinase (GSK)-3β, p-extracellular signal-regulated kinases (p-ERK), and p-(protein kinase B) AKT. Results: The in vitro results revealed that wogonoside exerted anti-proliferative activity against bladder cancer 5637 cells with an IC50 of 20.59 μM (p < 0.01), in a concentration- and time-dependent manner. Furthermore, wogonoside treatment also significantly suppressed tumor volume in mice (p < 0.01). The potential mechanisms were mainly associated with apoptosis mediated by mitochondria via upregulation of caspase-3, caspase-9, and Bax levels and down-regulation of Bcl-2, p-GSK-3β, p-ERK, and p-AKT. Conclusion: The results reveal that wogonoside has remarkable anti-tumor potentials against bladder cancer. Further translational studies are warranted to test the clinical application of this medicinal agent in bladder cancer.

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