Objective: To explore the effect of lncRNA ADPGK-AS1 on the proliferation and apoptosis of retinoblastoma cells and its possible mechanism. Methods: The tumor tissues of 31 patients with retinoblastoma admitted to Henan Provincial Eye Hospital from February to June 2020 and their corresponding normal tissues adjacent to the cancer were collected. The expression levels of lncRNA ADPGK-AS1 and miR-200b-5p in retinoblastoma tissues and normal adjacent tissues were detected by real-time fluorescence quantitative polymerase chain reaction (qRT-PCR). Human retinal epithelial cell ARPE-19, human retinoblastoma cell Y-79 and WERI-Rb-1 were cultured in vitro. The expression levels of lncRNA ADPGK-AS1 and miR-200b-5p were detected by qRT-PCR. Y-79 cells were randomly divided into si-con group, si-lncRNA ADPGK-AS1 group, miR con group, miR-200b-5p group, si-lncRNA ADPGK-AS1+ anti-miR con group, and si-lncRNA ADPGK-AS1+ anti-miR-200b-5p group. The proliferation, cloning and apoptosis of cells in each group were detected by tetramethylazol blue method, plate cloning test and flow cytometry, respectively. The targeting relationship between lncRNA ADPGK-AS1 and miR-200b-5p was detected by double luciferase report test, and the expression level of cleaved-caspase-3 protein was detected by western blot. Results: Compared with the adjacent tissues, the expression of lncRNA ADPGK-AS1 in retinoblastoma tissues was increased (P<0.05), while the expression of miR-200b-5p was decreased (P<0.05). Compared with ARPE-19 cells, the expression of lncRNA ADPGK-AS1 in Y-79 and WERI-Rb-1 cells was increased (P<0.05), while the expression of miR-200b-5p was decreased (P<0.05). Compared with the si-con group, the cell viability of the si-lncRNA ADPGK-AS1 group was reduced (1.06±0.09 vs 0.53±0.05, P<0.05), the number of cell clone formation was reduced (114.00±8.03 vs 57.00±4.13, P<0.05), while the apoptosis rate [(7.93±0.68)% vs (25.43±1.94)%] and the protein level of cleaved-caspase-3 were increased (P<0.05). Compared with the miR-con group, the cell viability of the miR-200b-5p group was decreased (1.05±0.08 vs 0.57±0.05, P<0.05), the number of cell clone formation was decreased (118.00±10.02 vs 64.00±5.13, P<0.05), while the apoptosis rate [(7.89±0.71)% vs (23.15±1.62)%] and the protein level of cleaved-caspase-3 were increased (P<0.05). lncRNA ADPGK-AS1 could target the expression of miR-200b-5p. Compared with the si-lncRNA ADPGK-AS1+ anti-miR-con group, cell viability of the si-lncRNA ADPGK-AS1+ anti-miR-200b-5p group was increased (0.53±0.04 vs 1.25±0.10, P<0.05), and the number of cell clones was increased (54.00±4.39 vs 125.00±10.03, P<0.05), while the rate of apoptosis [(25.38±1.53)% vs (9.76±0.71)%] and the protein level of cleaved-caspase-3 were decreased (P<0.05). Conclusion: Interfering with the expression of lncRNA ADPGK-AS1 could inhibit the proliferation and clone formation and induce apoptosis of retinoblastoma cells by targeting the expression of miR-200b-5p.
Humans ; MicroRNAs/metabolism* ; Retinoblastoma/pathology* ; Caspase 3/metabolism* ; RNA, Long Noncoding/metabolism* ; Antagomirs/pharmacology* ; Cell Proliferation ; Cell Line, Tumor ; Apoptosis/genetics* ; Retinal Neoplasms/genetics* ; Gene Expression Regulation, Neoplastic ; Cell Movement/genetics*

This study explores the effect of total flavonoids of Rhododendra simsii(TFR) on middle cerebral artery occlusion(MCAO)-induced cerebral injury in rats and oxygen-glucose deprivation/reoxygenation(OGD/R) injury in PC12 cells and the underlying mechanism. The MCAO method was used to induce focal ischemic cerebral injury in rats. Male SD rats were randomized into sham group, model group, and TFR group. After MCAO, TFR(60 mg·kg~(-1)) was administered for 3 days. The content of tumor necrosis factor-α(TNF-α), interleukin-1(IL-1), and interleukin-6(IL-6) in serum was detected by enzyme-linked immunosorbent assay(ELISA). The pathological changes of brain tissue and cerebral infarction were observed based on hematoxylin and eosin(HE) staining and 2,3,5-triphenyltetrazolium chloride(TTC) staining. RT-qPCR and Western blot were used to detect the mRNA and protein levels of calcium release-activated calcium channel modulator 1(ORAI1), stromal interaction molecule 1(STIM1), stromal intera-ction molecule 2(STIM2), protein kinase B(PKB), and cysteinyl aspartate specific proteinase 3(caspase-3) in brain tissues. The OGD/R method was employed to induce injury in PC12 cells. Cells were randomized into the normal group, model group, gene silencing group, TFR(30 μg·mL~(-1)) group, and TFR(30 μg·mL~(-1))+gene overexpression plasmid group. Intracellular Ca~(2+) concentration and apoptosis rate of PC12 cells were measured by laser scanning confocal microscopy and flow cytometry. The effect of STIM-ORAI-regulated store-operated calcium entry(SOCE) pathway on TFR was explored based on gene silencing and gene overexpression techniques. The results showed that TFR significantly alleviated the histopathological damage of brains in MCAO rats after 3 days of admini-stration, reduced the contents of TNF-α, IL-1, and IL-6 in the serum, down-regulated the expression of ORAI1, STIM1, STIM2, and caspase-3 genes, and up-regulated the expression of PKB gene in brain tissues of MCAO rats. TFR significantly decreased OGD/R induced Ca~(2+) overload and apoptosis in PC12 cells. However, it induced TFR-like effect by ORAI1, STIM1 and STIM2 genes silencing. However, overexpression of these genes significantly blocked the effect of TFR in reducing Ca~(2+) overload and apoptosis in PC12 cells. In summary, in the early stage of focal cerebral ischemia-reperfusion injury and OGD/R-induced injury in PC12 cells TFR attenuates ischemic brain injury by inhibiting the STIM-ORAI-regulated SOCE pathway and reducing Ca~(2+) overload and inflammatory factor expression, and apoptosis.
Animals ; Male ; Rats ; Apoptosis ; Brain Ischemia/metabolism* ; Caspase 3 ; Interleukin-1 ; Interleukin-6 ; Rats, Sprague-Dawley ; Reperfusion Injury/metabolism* ; Tumor Necrosis Factor-alpha/genetics* ; Flavonoids/pharmacology* ; Rhododendron/chemistry*

The aim of this study was to explore the effects of Huangqin Tang(HQT) on the NLRP3/Caspase-1 signaling pathway in mice with DSS-induced ulcerative colitis(UC). C57BL/6J mice were randomly divided into a blank group, a model group(DSS group), and low-, medium-and high-dose HQT groups(HQT-L, HQT-M, and HQT-H), and western medicine mesalazine group(western medicine group). The UC model was induced in mice. Subsequently, the mice in the HQT-L, HQT-M, HQT-H groups, and the western medicine group were given low-, medium-, high-dose HQT, and mesalazine suspension by gavage, respectively, while those in the blank and DSS groups were given an equal volume of distilled water by gavage. After 10 days of administration, the body weight, DAI scores, and colonic histopathological score of mice in each group were determined. The levels of IL-6, IL-10, IL-1β, and TNF-α in serum were determined by ELISA. The mRNA expression of NLRP3 and Caspase-1 in colon tissues was determined by RT-qPCR. The protein expression of NLRP3 and Caspase-1 in colon tissues was detected by immunohistochemistry. The results showed that compared with the blank group, the DSS group showed decreased body weight of mice and increased DAI scores and intestinal histopathological score. Compared with the DSS group, the HQT groups and the western medicine group showed improved DAI scores, especially in the HQT-M, HQT-H, and the western medicine groups(P<0.05). The intestinal histopathological scores of the HQT groups and the western medicine group significantly decreased, especially in the HQT-M, HQT-H, and the western medicine groups(P<0.05). In addition, compared with the blank group, the DSS group showed elevated expression of NLRP3 and Caspase-1 in colon tissues, increased serum levels of IL-6, IL-1β, and TNF-α, and decreased IL-10 level. Compared with the DSS group, the HQT groups and the western medicine group displayed decreased expression of NLRP3 and Caspase-1 in colon tissues, reduced serum levels of IL-6, IL-1β, and TNF-α, and increased IL-10 level. The improvement was the most significant in the HQT-H group and the western medicine group(P<0.01). In conclusion, HQT may reduce the expression of NLRP3 and Caspase-1 in colon tissues, reduce the se-rum levels of IL-6, IL-1β, and TNF-α, and increase the expression of IL-10 by regulating the classic pyroptosis pathway of NLRP3/Caspase-1, thereby improving the symptoms of intestinal injury and inflammatory infiltration of intestinal mucosa in DSS mice to achieve its therapeutic effect.
Animals ; Mice ; Caspase 1/genetics* ; Colitis, Ulcerative/genetics* ; Colon ; Dextran Sulfate/adverse effects* ; Disease Models, Animal ; Interleukin-10/genetics* ; Interleukin-6/genetics* ; Mesalamine/pharmacology* ; Mice, Inbred C57BL ; NLR Family, Pyrin Domain-Containing 3 Protein/genetics* ; Scutellaria baicalensis/chemistry* ; Tumor Necrosis Factor-alpha/metabolism* ; Drugs, Chinese Herbal/pharmacology*

Objective: To investigate the potential function and related mechanism of microRNA-223 (miRNA-223) in the podocyte pyroptosis of hepatitis B virus (HBV)-associated glomerulonephritis induced by HBV X protein (HBx). Methods: HBx-overexpressing lentivirus was transfected into human renal podocytes to mimic the pathogenesis of HBV-GN. Real-time fluorescence quantitative PCR and Western blotting experiments were used to detect the mRNA and protein expression of pyroptosis-related proteins [nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3), apoptosis-associated speck-like protein containing a CARD (ASC) and caspase-1], and inflammatory factors (interleukin-1β and interleukin-18), respectively.TUNEL staining and flow cytometry were used to detect the number of pyroptosis cells. Immunofluorescence staining was used to detect the expression of podocytes biomarkers desmin and nephrin; Hoechst 33342 staining was used to observe the morphological and quantitative changes of podocyte nuclei. Enzyme-linked immunosorbent assay was used to measure caspase-1 activity. The dual luciferase reporter gene assay was used to verify the downstream target of miRNA-223. Podocytes were divided into the following nine groups: control group (no special treatment), empty plasmid group (transfected with empty plasmid), HBx overexpression group (transfected with HBx overexpression lentivirus), HBx overexpression+miRNA-223 mimic group (transfected with HBx overexpression lentivirus and miRNA-223 mimic), HBx overexpression+miRNA-223 inhibitor group (transfected with HBx overexpression lentivirus and miRNA-223 inhibitor), HBx overexpression+miRNA-223 mimic+NLRP3 group (transfected with HBx overexpression lentivirus, miRNA-223 mimic and NLRP3 overexpression plasmid), HBx overexpression+miRNA-223 mimic+ NLRP3 siRNA group (transfected with HBx overexpression lentivirus, miRNA-223 mimic and NLRP3 siRNA), HBx overexpression+miRNA-223 inhibitor+NLRP3 group (transfected with HBx overexpression lentivirus, miRNA-223 inhibitor and NLRP3 overexpression plasmid), HBx overexpression+miRNA-223 inhibitor+NLRP3 siRNA group (transfected with HBx overexpression lentivirus, miRNA-223 inhibitor and NLRP3 siRNA). Results: miRNA-223 was down-regulated in HBx overexpression group compared with the control group (P < 0.05). TUNEL and immunofluorescence staining showed that NLRP3 knockdown attenuated podocyte injury and pyroptosis induced by HBx overexpression (P < 0.05). Dual luciferase reporter gene assay demonstrated that NLRP3 was one of the downstream targets of miRNA-223. Rescue experiments revealed that NLRP3 overexpression weakened the protective effect of miRNA-223 in podocyte injury (P < 0.05). The addition of miRNA-223 mimic and NLRP3 siRNA decreased the expression of NLRP3 inflammasome and cytokines, and reduced the number of pyroptosis cells induced by HBx overexpression (all P < 0.05); The addition of miRNA-223 inhibitor and NLRP3 overexpression plasmid significantly increased the expression of NLRP3 inflammasome and cytokines, caspase-1 activity, and the number of pyroptosis cells (all P < 0.05). Conclusion: HBx may promote podocyte pyroptosis of HBV-GN via downregulating miRNA-223 targeting NLRP3 inflammasome, suggesting that miRNA-223 is expected to be a potential target for the treatment of HBV-GN.
Humans ; Inflammasomes/metabolism* ; NLR Family, Pyrin Domain-Containing 3 Protein/metabolism* ; Pyroptosis ; Podocytes/metabolism* ; Hepatitis B virus/genetics* ; Caspase 1/metabolism* ; Cytokines/metabolism* ; Carrier Proteins/metabolism* ; MicroRNAs/genetics* ; Glomerulonephritis/metabolism* ; RNA, Small Interfering

OBJECTIVE: To explore the effect of abnormal miRNA expression on the proliferation of pediatric acute lymphoblastic leukemia (ALL) cells and its related mechanism. METHODS: 15 children with ALL and 15 healthy subjects were collected from the Second Affiliated Hospital of Hainan Medical University from July 2018 to March 2021. MiRNA sequencing was performed on their bone marrow cells, and validated using qRT-PCR. MiR-1294 and miR-1294-inhibitory molecule (miR-1294-inhibitor) were transfected into Nalm-6 cells, and the proliferation of Nalm-6 cells was detected by CCK-8 and colony formation assays. Western blot and ELISA were used to detect apoptosis of Nalm-6 cells. Biological prediction of miR-1294 was performed to find the target gene, which was verified by luciferase reporter assay. Si-SOX15 was transfected into Nalm-6 cells, Western blot was used to detect the expression of Wnt signaling pathway-related proteins and to verify the effect of si-SOX15 on the proliferation and apoptosis of Nalm-6 cells. RESULTS: Compared with healthy subjects, 22 miRNAs were significantly upregulated in bone marrow cells of ALL patients, of which miR-1294 was the most significantly upregulated. In addition, the expression level of SOX15 gene was significantly reduced in bone marrow cells of ALL patients. Compared with the NC group, the miR-1294 group showed increased protein expression levels of Wnt3a and β-catenin, faster cell proliferation, and more colony-forming units, while caspase-3 protein expression level and cell apoptosis were reduced. Compared with the NC group, the miR-1294-inhibitor group showed reduced protein expression levels of Wnt3a and β-catenin, slower cell proliferation, and fewer colony-forming units, while caspase-3 protein expression level was increased and apoptosis rate was elevated. miR-1294 had a complementary base-pair with the 3'UTR region of SOX15 , and miR-1294 directly targeted SOX15 . The expression of miR-1294 was negatively correlated with SOX15 in ALL cells. Compared with the si-NC group, the si-SOX15 group showed increased protein expression levels of Wnt3a and β-catenin, accelerated cell proliferation, and decreased caspase-3 protein expression level and cell apoptosis rate. CONCLUSION MiR-1294 can target and inhibit SOX15 expression, thus activating the Wnt/β-Catenin signaling pathway to promote the proliferation of ALL cells, inhibit cell apoptosis, and ultimately affect the disease progression.
Humans ; Child ; beta Catenin/genetics* ; Wnt Signaling Pathway ; Caspase 3/metabolism* ; Cell Line, Tumor ; MicroRNAs/genetics* ; Cell Proliferation ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; Apoptosis ; SOX Transcription Factors/metabolism*

Objective: To investigate the effects of lncRNA DRAIC on proliferation, apoptosis, migration and invasion of lung adenocarcinoma cells and its mechanism. Methods: Reverse transcription-quantitative real-time polymerase chain reaction (RT-qPCR) was used to detect the expression of DRAIC in lung cancer tissues and corresponding adjacent normal tissues of 40 patients with lung adenocarcinoma who underwent surgery in Tangshan People's Hospital from 2019 to 2020. Lung adenocarcinoma cells A549 and H1299 were cultured in vitro and divided into si-NC group, si-DRAIC group, miR-NC group, let-7i-5p mimics group, si-DRAIC+ inhibitor-NC group, and si-DRAIC+ let-7i-5p inhibitor group. CCK-8 method and clone formation experiment were used to detect cell proliferation. Flow cytometry was used to detect cell apoptosis. Transwell array was used to detect the cell migration and invasion. Western blot was used to detect the protein expressions of Caspase-3, Caspase-9, Bcl-2 and Bax. The double luciferase reporter gene experiment was used to verify the regulatory relationship between DRAIC and let-7i-5p. Independent sample t test was used for comparison between two groups, one-way ANOVA was used for comparison between multiple groups, and Pearson correlation analysis was used for correlation analysis. Results: Compared with adjacent tissues, the expression level of DRAIC in lung adenocarcinoma tissues increased (P<0.05), but the expression level of let-7i-5p decreased (P<0.05). The expression levels of DRAIC and let-7i-5p in lung adenocarcinoma tissues were negatively correlated (r=-0.737, P<0.05). The absorbance value of A549 and H1299 cells in the si-DRAIC group at 48, 72 and 96 hours were lower than those in the si-NC group (P<0.05), the number of clones formed [(91.00±6.08 vs. 136.67±6.51); (50.67±1.53 vs. 76.67±4.51)], the number of migration [(606.67±31.34 vs. 960.00±33.06); (483.33±45.96 vs. 741.67±29.67)], the number of invasion [(185.00±8.19 vs. 447.33±22.05); (365.00±33.87 vs. 688.00±32.97)] were lower than those in the si-NC group (P<0.05). However, the apoptosis rates of cells [(13.43±2.79)% vs. (4.53±0.42)%; (23.77±1.04)% vs. (6.60±1.42)%] were higher than those in the si-NC group (P<0.05). The protein expressions of Caspase-3, Caspase-9 and Bax in si-DRAIC group were higher than those in si-NC group, and the protein expression of Bcl-2 was lower than that in si-NC group (P<0.05). DRAIC is located in the cytoplasm. DRAIC targeted and negatively regulated the expression of let-7i-5p. The absorbance values of A549 and H1299 cells in the let-7i-5p mimics group at 48, 72 and 96 hours were lower than those in the miR-NC group (P<0.05), the number of clones formed [(131.33±14.47 vs. 171.33±6.11); (59.33±4.93 vs. 80.33±7.09)], the number of migration [(137.67±3.06 vs. 579.33±82.03); (425.00±11.14 vs. 669.33±21.13)], the number of invasion [(54.00±4.36 vs. 112.67±11.59); (80.00±4.58 vs. 333.33±16.80)] were lower than those in the miR-NC group (P<0.05). However, the apoptosis rates of cells [(14.57±1.10)% vs. (6.97±1.11)%; (23.97±0.42)% vs. (7.07±1.21)%] were higher than those in the miR-NC group (P<0.05). The protein expressions of Caspase-3, Caspase-9 and Bax in let-7i-5p mimics group were higher than those in miR-NC group, and the protein expression of Bcl-2 was lower than that in miR-NC group (P<0.05). The absorbance values of A549 and H1299 cells in the si-DRAIC+ let-7i-5p inhibitor group at 48, 72 and 96 hours were higher than those in the si-DRAIC+ inhibitor-NC group (P<0.05), the number of clones formed [(82.00±5.29 vs. 59.00±5.57); (77.67±4.93 vs. 41.33±7.57)], the number of migration [(774.33±35.81 vs. 455.67±19.04); (569.67±18.72 vs. 433.67±16.77)], the number of invasion [(670.33±17.21 vs. 451.00±17.52); (263.67±3.06 vs. 182.33±11.93)] were higher than those in the si-DRAIC+ inhibitor-NC group (P<0.05). However, the apoptosis rates of cells [(7.73±0.45)% vs. (19.13±1.50)%; (8.00±0.53)% vs. (28.40±0.53)%] were lower than those in the si-NC group (P<0.05). The protein expressions of Caspase-3, Caspase-9 and Bax in si-DRAIC+ let-7i-5p inhibitor group were higher than those in si-DRAIC+ inhibitor-NC group, and the protein expression of Bcl-2 was lower than that in si-DRAIC+ inhibitor-NC group (P<0.05). Conclusion: DRAIC is highly expressed in lung adenocarcinoma, and DRAIC promotes the proliferation, migration and invasion of lung adenocarcinoma cells and inhibits apoptosis by targeting let-7i-5p.
Humans ; Adenocarcinoma/genetics* ; Apoptosis/genetics* ; bcl-2-Associated X Protein/metabolism* ; Caspase 3/metabolism* ; Caspase 9/metabolism* ; Cell Line, Tumor ; Cell Proliferation/genetics* ; Lung/metabolism* ; MicroRNAs/metabolism* ; Proto-Oncogene Proteins c-bcl-2/metabolism* ; RNA, Long Noncoding/genetics*

OBJECTIVE: To investigate the expression of microRNA miR-431-5p in gastric cancer (GC) tissues and its effects on apoptosis and mitochondrial function in GC cells. METHODS: The expression level of miR-431-5p in 50 clinical samples of GC tissues and paired adjacent tissues was detected using real-time fluorescence quantitative PCR, and its correlation with the clinicopathological features of the patients was analyzed. A cultured human GC cell line (MKN-45 cells) were transfected with a miR-431-5p mimic or a negative control sequence, and the cell proliferation, apoptosis, mitochondrial number, mitochondrial potential, mitochondrial permeability transition pore (mPTP), reactive oxygen species (ROS) production and adenosine triphosphate (ATP) content were detected using CCK-8 assay, flow cytometry, fluorescent probe label, or ATP detection kit. The changes in the expression levels of the apoptotic proteins in the cells were detected with Western blotting. RESULTS: The expression level of miR-431-5p was significantly lower in GC tissues than in the adjacent tissues (P < 0.001) and was significantly correlated with tumor differentiation (P=0.0227), T stage (P=0.0184), N stage (P=0.0005), TNM stage (P=0.0414) and vascular invasion (P=0.0107). In MKN-45 cells, overexpression of miR-431-5p obviously inhibited cell proliferation and induced cell apoptosis, causing also mitochondrial function impairment as shown by reduced mitochondrial number, lowered mitochondrial potential, increased mPTP opening, increased ROS production and reduced ATP content. Overexpression of miR-431-5p significantly downregulated the expression of Bcl-2 and increased the expressions of pro-apoptotic proteins p53, Bcl-2 and cleaved caspase-3 protein. CONCLUSION The expression of miR-431-5p is down-regulated in GC, which results in mitochondrial function impairment and promotes cell apoptosis by activating the Bax/Bcl-2/caspase3 signaling pathway, suggesting the potential role of miR-431-5p in targeted therapy for GC.
Humans ; Apoptosis/genetics* ; bcl-2-Associated X Protein ; Caspase 3 ; Cell Line, Tumor ; Cell Proliferation/genetics* ; MicroRNAs/metabolism* ; Mitochondria/metabolism* ; Mitochondrial Permeability Transition Pore ; Reactive Oxygen Species ; Stomach Neoplasms/pathology*

Objective To explore the effect of knocking down Rho-associated coiled-coil kinase (ROCK2) gene on the cognitive function of amyloid precursor protein/presenilin-1 (APP/PS1) double transgenic mice and its mechanism. Methods APP/PS1 double transgenic mice were randomly divided into AD model group (AD group), ROCK2 gene knock-down group (shROCK2 group), ROCK2 gene knock-down control group (shNCgroup), and wild-type C57BL/6 mice of the same age served as the wild-type control (WT group). Morris water maze and Y maze were employed to test the cognitive function of mice. Neuron morphology was detected by Nissl staining. Immunofluorescence histochemical staining was used to detect the expression of phosphorylated dynamin-related protein 1 (p-Drp1) and mitochondrial fusion 1 (Mfn1). Western blot analysis was used to detect the expression ROCK2, cleaved-caspase-3 (c-caspase-3), B-cell lymphoma 2 (Bcl2), Bcl2-related protein X (BAX), p-Drp1, mitochondrial fission 1 (Fis1), optic atrophy 1 (OPA1), Mfn1 and Mfn2. Results Compared with AD group mice, the expression of ROCK2 in shROCK2 group mice was significantly reduced; the cognitive function was significantly improved with the number of neurons in the hippocampal CA3 and DG areas increasing, and nissl bodies were deeply stained; the expression of c-caspase-3 and BAX was decreased, while the expression of Bcl2 was increased; the expression of mitochondrial division related proteins p-Drp1 and Fis1 were decreased, while the expression of mitochondrial fusion-related proteins OPA1, Mfn1 and Mfn2 were increased. Conclusion Knock-down of ROCK2 gene can significantly improve the cognitive function and inhibit the apoptosis of nerve cells of APP/PS1 mice. The mechanism may be related to promoting mitochondrial fusion and inhibiting its division.
Animals ; Mice ; Alzheimer Disease/pathology* ; Amyloid beta-Peptides/metabolism* ; Amyloid beta-Protein Precursor ; Apoptosis/genetics* ; bcl-2-Associated X Protein ; Caspase 3 ; Cognition ; Disease Models, Animal ; Mice, Inbred C57BL ; Mice, Transgenic ; Mitochondrial Dynamics/genetics*

OBJECTIVE: To investigate the effect of long non-coding RNA LINC01268 on apoptosis of acute myeloid leukemia (AML) cells and related mechanisms. METHODS: The expression levels of LINC01268 and miR-217 in peripheral blood samples from AML patients and AML cell lines HL-60 and KG-1 were detected by qRT-PCR. HL-60 cells were divided into pcDNA3.1-NC, pcDNA3.1-LINC01268, si-NC, si-LINC01268, miR-NC, miR-217 mimics, si-LINC01268 + inhibitor-NC and si-LINC01268+ miR-217 inhibitor groups. The mRNA expressions of LINC01268 and miR-217 were detected by qRT-PCR. The targeting relationship between LINC01268 and miR-217 was detected by dual-luciferase reporter assay. Cell viability was detected by CCK-8 assay. Cell cycle distribution and apoptosis were detected by flow cytometry. The expression of cell cycle and apoptosis-related proteins p21, Bcl-2, Bax, caspase-3 and PI3K/AKT signaling pathway-related proteins were detected by Western blot. RESULTS: The expression of LINC01268 in peripheral blood samples of AML patients and AML cell lines HL-60 and KG-1 was increased (P < 0.05), and the expression of miR-217 was decreased (P < 0.05). Compared with si-NC group and miR-NC group, the viability of HL-60 cells was decreased in si-LINC01268 group and miR-217 mimics group (P < 0.05), the proportion of cells in G₁ phase and apoptosis rate were increased (P < 0.05), the protein expression levels of p21, Bax and caspase-3 were increased (P < 0.05), while the protein expression level of Bcl-2 was decreased (P < 0.05). LINC01268 targeted and negatively regulated the expression of miR-217, and inhibiting the expression of miR-217 partially reversed the effects of LINC01268 interference on the viability, cell cycle and apoptosis of HL-60 cells. Interference with LINC01268 could inhibit the activity of PI3K/AKT signaling pathway. Inhibiting the expression of miR-217 could partially reverse the inhibition of LINC01268 interference on PI3K/AKT signaling pathway. CONCLUSION LINC01268 is highly expressed and miR-217 is lowly expressed in AML cells. LINC01268 can promote the activity of PI3K/AKT signaling pathway, increase the survival rate and inhibit the apoptosis of AML cells by targeting miR-217 expression.
Humans ; Apoptosis ; bcl-2-Associated X Protein/metabolism* ; Caspase 3 ; Cell Line, Tumor ; Cell Proliferation ; Leukemia, Myeloid, Acute/metabolism* ; MicroRNAs/metabolism* ; Phosphatidylinositol 3-Kinases/metabolism* ; Proto-Oncogene Proteins c-akt/metabolism* ; RNA, Long Noncoding/genetics*

To investigate the antidepressant mechanism of Shenling Kaixin Granules(SLKX) in treating chronic unpredictable mild stress(CUMS) model rats. Ninety male SD rats were randomly divided into control group, model group, Shugan Jieyu Capsules(110 mg·kg~(-1)) group and SLKX low-(90 mg·kg~(-1)), medium-(180 mg·kg~(-1)), and high-dose(360 mg·kg~(-1)) groups. Depression rat model was replicated by CUMS method. After treatment, the behavioral changes of rats were evaluated by sugar preference, open field, elevated cross maze and forced swimming experiments. The contents of interleukin 1 beta(IL-1β), tumor necrosis factor α(TNF-α), brain-derived neurotrophic factor(BDNF) and 5-hydroxytryptamine(5-HT) in serum were determined by enzyme linked immunosorbent assay(ELISA), and the activities of superoxide dismutase(SOD) and catalase(CAT) in hippocampal CA1 region were also detected. Pathological changes in hippocampal CA1 region were detected by hematoxylin-eosin(HE) staining, and Western blot was used to determine the expression of nerve growth factor(NGF), BDNF, phospho-tyrosine kinase receptor(p-TrkB)/TrkB, phospho-cAMP-response element binding protein(p-CREB)/CREB, nuclear factor E2 related factor 2(Nrf2), heme oxygenase 1(HO-1), B-cell lymphoma-2(Bcl-2)/Bcl-2 associated X protein(Bax) and caspase-3 in hippocampal CA1 region. RESULTS:: showed that compared with the control group, the model group had decreased sugar preference, reduced number of entries and time spent in the center of open field and shortened total distance of movement, reduced number of entries and proportion of time spent in open arm, and increased number and time of immobility in forced swimming experiment. Additionally, the serum contents of IL-1β and TNF-α and the expression of caspase-3 were higher, while the contents of BDNF and 5-HT, the activities of SOD and CAT in hippocampal CA1 region, the expressions of NGF, BDNF, p-TrkB/TrkB, p-CREB/CREB, HO-1 and Bcl-2/Bax, and the Nrf2 nuclear translocation were lower in model group than in control group. Compared with the conditions in model group, the sugar preference, the number of entries and time spent in the center of open, total distance of movement, and the number of entries and proportion of time spent in open arm in treatment groups were increased while the number and time of immobility in forced swimming experiment were decreased; the serum contents of IL-1β and TNF-α and the expression of caspase-3 were down regulated, while the contents of BDNF and 5-HT, the activities of SOD and CAT in hippocampal CA1 region, the expressions of NGF, BDNF, p-TrkB/TrkB, p-CREB/CREB, HO-1, Bcl-2/Bax, and Nrf2 nuclear translocation were enhanced. In conclusion, SLKX might regulate the Nrf2 nucleus translocation by activating BDNF/TrkB/CREB pathway, lower oxidative stress damage in hippocampus, inhibit caspase-3 activity, and reduce apoptosis of hippocampal nerve cells, thereby playing an antidepressant role.
Rats ; Male ; Animals ; bcl-2-Associated X Protein/metabolism* ; Caspase 3/metabolism* ; Nerve Growth Factor/metabolism* ; Brain-Derived Neurotrophic Factor/metabolism* ; Signal Transduction ; Tumor Necrosis Factor-alpha/metabolism* ; Serotonin/metabolism* ; NF-E2-Related Factor 2/metabolism* ; Rats, Sprague-Dawley ; Antidepressive Agents/pharmacology* ; Hippocampus/metabolism* ; Superoxide Dismutase/metabolism* ; Sugars/pharmacology* ; Depression/genetics* ; Stress, Psychological/metabolism*