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Objective To investigate the effect of anti-angiogenic drug Sitravatinib combined with poly(adenosine diphosphate[ADP]-ribose)polymerase inhibitor(PARPi)Niraparib on mucosal melanoma cell lines and its possible mechanism.Methods The CCK8 assay was used to detect the maximal half inhibitory concentration(IC50)of Sitravatinib and Niraparib targeting at mucosal melanoma(MM)cell lines.CompuSyn was used to detect the Combination Index(CI)in different concentrations of the two drugs.Flow cytometry was used to detect the effect of drugs on cell apoptosis.Colony formation assay was used to detect the effect of drugs on cell proliferation.Western blot was used to detect the protein expressions and RT-qPCR was used to detect mRNA expression.Results CI values was respectively 0.19 and 0.15 for Sitravatinib(2 μmol/L)in combination with Niraparib(20 μmol/L)in a human vaginal maligant melanoma cell line(HMVII)and a metastasis inguinal lymph node of vulvar malignant melanoma cell line(GAK).Compared with the control group and single-drug groups,the cell proliferation of the combination group was significantly reduced(P＜0.05 or P＜0.01 or P＜0.001).The cell apoptosis rate was signifi-cantly increased(P＜0.01 or P＜0.001).The protein and mRNA expression of apoptosis-related biomarkers signifi-cantly increased(P＜0.001);In addition,the protein and mRNA expression of cell autophagy biomarkers signifi-cantly increased(P＜0.01 or P＜0.001).The protein expression of DNA damage marker significantly increased.Moreover,compared with the control group,The expression of radiation sensitive protein 51(RAD51)recombinase in the Sitravatinib single-drug group and combination group significantly reduced.As the dose of Sitravatinib gradu-ally increased up to 2 μmol/L,the protein and mRNA expression of RAD51 both significantly reduced(P＜0.05 or P＜0.01),the mRNA expression of BRCA1 and BRCA2 also significantly reduced(P＜0.05 or P＜0.01 or P＜0.001).Conclusions Sitravatinib combined with Niraparib inhibits the proliferation of mucosal melanoma cells,induces cell apoptosis and promotes autophagy.The mechanism is potentially related to the inhibition of ho-mology-dependent recombination repairs(HRR).
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Objective To investigate the protective effect of human umbilical cord mesenchymal stem cell-derived exosome (hucMSC-Exo) on renal ischemia-reperfusion injury (IRI), and to clarify the critical role and regulating mechanism of transient receptor potential canonical (TRPC) 6/poly adenosine-diphosphate-ribose polymerase (PARP) 1 signaling pathway during this process. Methods The hucMSC-Exo was extracted by ultracentrifugation, and identified by transmission electron microscope (TEM), nanoparticle tracing analysis and Western blot. SD rats were randomly divided into the sham operation group (group S), sham operation+TRPC6 inhibitor SKF96365 group (group SS), renal IRI group (group IRI), exosome treatment group (group EXO) and exosome +TRPC6 inhibitor SKF96365 group (group ES), with 6 rats in each group. Serum creatinine and blood urea nitrogen levels were detected. Pathological changes of renal tissues were observed by hematoxylin-eosin (HE) staining and Paller score was calculated. The expression levels of key molecules of necroptosis in rat renal tissues, including receptor-interacting protein kinase (RIPK)1, RIPK3 and mixed-lineage kinase domain-like protein (MLKL), TRPC6 and PARP1, were detected by Western blot. Results Typical saucer-like structure was observed under TEM. Nanoparticle tracing analysis showed that the average diameter of the extracted substance was 125.9 nm. Western blot revealed that the surface markers of CD9, CD63 and CD81 were positively expressed, confirmed that the extracted substance was exosome. Compared with group S, the serum creatinine and blood urea nitrogen levels were up-regulated, the pathological damage of renal tissues was worsened, Paller score was elevated, the relative expression levels of TRPC6 and PARP1 proteins were down-regulated, and the relative expression levels of RIPK1, RIPK3 and MLKL proteins were up-regulated in group IRI (all P < 0.05). Compared with group IRI, the serum creatinine and blood urea nitrogen levels were down-regulated, the pathological damage of renal tissues was mitigated, Paller score was decreased, the relative expression levels of TRPC6 and PARP1 proteins were up-regulated, and the relative expression levels of RIPK1, RIPK3 and MLKL proteins were down-regulated in group EXO (all P < 0.05). Compared with group EXO, the serum creatinine and blood urea nitrogen levels were up-regulated, the pathological damage of renal tissues was aggravated, Paller score was increased, the relative expression levels of TRPC6 and PARP1 proteins were down-regulated, and the relative expression levels of RIPK1, RIPK3 and MLKL proteins were up-regulated in group ES (all P < 0.05). Conclusions hucMSC-Exo may alleviate the necroptosis induced by renal IRI in rat models, which is related to the activation of TRPC6/PARP1 signaling pathway.
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Objective:To investigate the effects of poly adenosine diphosphate ribose polymerase-1(PARP-1) inhibitor fluzoparib on proliferation, apoptosis and migration of pancreatic cancer PANC1 cells.Methods:PANC1 cells cultured in conventional culture medium were used as control group, and PANC1 cells cultured in the medium containing fluzoparib were used as fluzoparib group. The effects of fluzoparib with different concentrations on the proliferation of PANC1 cells were detected by CCK8 method, and the half inhibitory concentration (IC 50) of fluzoparib on PANC1 cells was calculated. The effect of fluzoparib on apoptosis and cell cycle of PANC1 cells was detected by flow cytometry, and the migration ability of PANC1 cells was detected by cell scratch test and Transwell chamber. Results:Compared with control group, with the increase of fluzoparib concentration and the prolongation of the action time, the cell proliferation activity of PANC1 in fluzoparib group was significantly decreased, and the differences were statistically significant (all P values <0.05). IC 50 of fluzoparib on PANC1 cells cultured for 24 h was 0.03 mmol/L. After 24 h culture, the IC 50 apoptosis rate of fluzoparib group was (32.19±2.48)%, and the apoptosis rate of control group was (21.99±6.30)%. The former was greatly higher than the latter, and the difference was statistically significant ( P<0.05). The proportion of cells in G 2/M phase was (16.28±0.62)% in the fluzoparib group and (11.64±0.88)% in the control group, and the difference between the two groups was statistically significant ( P<0.05). The migration rates of PANC1 cells in IC 50 fluzoparib group in 12 h and 24 h culture were (2.59±1.46)% and (19.76±7.84)%; and those in control group were (27.08±2.17)% and (45.92±3.61)%, respectively. The number of transmembrane cells was (348±19) cells/10 visual field in the fluzoparib group and (587±14) cells/10 visual field in the control group. The migration ability of PANC1 cells in fluzoparib group was significantly lower than that in control group ( P<0.05). Conclusions:Fluzoparib can inhibit the proliferation and migration of PANC1 cells and promote the apoptosis of PANC1 in vitro, which may be an effective drug for the treatment of pancreatic cancer.
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Ovarian cancer seriously threats women's health. The emergence of poly adenosine diphosphate ribose polymerase inhibitor (PARPi) has broken the barrier for ovarian cancer treatment. PARPi has been widely used and along with it comes the problem of drug resistance. Fully understanding the drug resistance mechanism of PARPi is expected to be an important way to reverse PARPi resistance. The combination of PARPi and other drugs for ovarian cancer may expand the benefits of patients using PARPi. This article reviews the drug resistance mechanism of PARPi and combined medication.
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Kidney ischemia-reperfusion injury (IRI) is the major cause of poor prognosis after kidney transplantation and partial nephrectomy. Besides, it is also a critical pathophysiological process of acute kidney injury. Consequently, the prevention and treatment of kidney IRI are of significance to improve clinical prognosis of recipients undergoing kidney transplantation. However, the mechanism underlying IRI is complex, and the exact mechanism remains elusive. Inflammation, as one of the main pathogenesis of IRI, plays a significant role in IRI-induced kidney injury. Nuclear factor (NF)-κB, as a rapid response transcription factor, has been proven to be involved in the regulation of inflammation during kidney IRI. Therefore, in this article, the structure of NF-κB, the activation pattern of NF-κB signaling pathway, the regulatory mechanisms of NF-κB upstream and downstream signaling pathways in kidney IRI were reviewed, and the role of NF-κB signaling pathway in kidney IRI was investigated, aiming to provide novel clinical ideas for the prevention and treatment of kidney IRI.
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Hereditary breast cancer refers to malignant tumors caused by pathogenic germline mutations of breast cancer susceptibility genes (BRCA). At present, it is believed that BRCA1/2 genes are most closely related to the development of hereditary breast cancer. Mutation will lead to loss of normal function, instability of genome, and then lead to tumorigenesis. Especially for those with germline mutations, not only the risk of breast cancer will be greatly increased, but also the probability of ovarian cancer and other cancers will be increased. With the emergence and clinical application of poly (adenosine diphosphate-ribose) polymerase (PARP) inhibitors, BRCA1/2 genes have been regarded as new targets for the treatment of breast cancer. This article reviews the latest research of breast cancer with BRCA1/2 gene mutations.
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Objective To investigate the effects of poly adenosine diphosphate ribose polymerase (PARP) inhibitor AG014699 on the proliferation of triple negative breast cancer (TNBC) cell line MDA?MB?231.Methods Cell proliferation and cytotoxicity test kit ( CCK?8) was used to detect the proliferation of MDA?MB?231 cells in different concentrations of AG014699 (0.1,1.0,10.0,20.0 and 40.0 mmol／L), DTX (10－9,10－8,10－7,10－6 and 10－5 mol／L) and CBP (10－6,10－5,10－4 and 10－3 mol／L).Flow cytometry was used to detect cell apoptosis and cell cycle distribution.Results The effects of AG01469 at different concentrations (0.1,1.0,10.0,20.0 and 40.0 μmol／L) on proliferation activity of MDA?MB?231 cells were (94.83 ± 3.93)%, ( 79.42 ± 5.52)%, ( 63.75 ± 4.34)%, ( 38.97 ± 8.42)%, ( 29.70 ± 3.35 )%, with statistically significant differences (F＝75.54,P＜0.01,different concentrations pairwise comparison: all P ＜0.05). The efficacy of AG014699 in combination with DTX at different concentrations (( 69.77 ±17.94)%,(58.34± 2.59)%,( 52.81 ± 2.01)%, ( 41.23 ± 3.38)%, ( 24.82 ± 0.73)%) was compared with that of single DTX (( 81.24 ± 11.91)%, ( 85.74 ± 3.10)%, ( 72.74 ± 4.66)%, ( 55.18 ± 3.19)%, (45.95±3.82)%).The differences were statistically significant (t values were －0.923,－11.748,－6.802,－5.199,－9.410,respectively,with P＞0.05 at 10－9 concentration and P＜0.01 at all other concentrations ). The efficacy of AG014699 combined with CBP ((78.33± 2.89)%,( 60.44± 1.95)%,( 50.55± 3.07)%, (12.07± 1.63)%) and single CBP (( 90.00 ± 6.18)%, ( 87.87 ± 2.30)%,( 76.82 ± 3.37)%,( 40.71 ±1.68)%) was compared,and the cell activity was significantly reduced,indicating statistically significant differences ( t values were －1.935,－15.756,－9.981,－21.192, respectively, and P＞0.05 at 10－6 concentration,P＜0.05 at all the other concentrations ).The q value was＞1.15 when AG014699 was combined with 10－3 mmol／L CBP, which showed synergistic effect.When combined with other effective concentrations of DTX or CBP,the q value was between 0.85 and 1.15,showing additive effect.Conclusion PARP inhibitor AG014699 assisted DTX or CBP can inhibit the proliferation of TNBC cell line MDA?MB?231.By means of simple addition or systematic effect,it can inhibit the triple negative breast cancer.
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Background Poly(adenosine diphosphate-ribose) polymerase 1 (PARP-1) plays an important role in the pathogenesis of diabetic retinopathy (DR),and Notch1 signal pathway is one of the important signal transduction pathways in the organism which may antagonize retinal vascular diseases.However,if Notch1 signaling pathway is involved in pathogenesis of DR has not been confirmed yet.Objective This study was to investigate the expressions of Notch1,Dll4,PARP-1,Akt,nuclear factor-κB (NF-κB) and caspase-3 in the retina of diabetic mouse model and retinal vascular endothelial cells (RVECs) under the high glucose.Methods The expressions of Notch1,Dll4,PARP-1,Akt,NF-κB and caspase-3 in the retina of diabetic mouse models were investigated using immunochemistry and Western blot method after the diabetic mouse models were established.And these proteins were detected in retinal RVECs under the high glucose by Western blot.Results The expressions of Notch1,Dll4 and phosphorylated Akt (p-Akt) in retinas reduced significantly and simultaneously companied with increases of PARP-1 and caspase-3 in diabetic mice compared with normal mice (all at P＜0.05).However,no obvious change was found in the expression of NF-κB (P＞0.05).Expressions of Notch1 and p-Akt in RVECs increased with the increase of glucose concentration,but expressions of cleaved PARP-1 and caspase-3 decreased,especially in the 30 mmol/L group,showing significant differences in comparison with the normal control group (all at P＜0.05).But no altering of NF-κB expression was seen in the mice with diabetes mellitus.Conclusions The expressions of cleaved-PARP-1 and caspase-3 in the retinas is up-regulated,but the expressions of Notch1 and p-Akt are down-regulated in diabetic mice.
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AIM:To investigate the time course of nuclear factor-κB (NF-κB) and the effects of 3-aminobenzamide (3-AB) on the expressions of NF-κB,interleukin-1β (IL-1β) and cyclooxygenase-2 (COX-2) in hippocampus after seizures. METHODS:Epilepsy were induced by kainic acid through cerebral ventricular injection. Western blotting was used to detect NF-κB p65 expression in nucleus at various experiment groups. Moreover,mRNA and protein expressions of IL-1β and COX-2 in different experiment groups were determined by RT-PCR and Western blotting analysis. RESULTS:NF-κB p65 immunoreactivity began to increase in the nuclear fraction at 2 h (P<0.05),kept rising at 12 h (P<0.05) and returned to control level at 24 h after epilepsy seizures. Furthermore,3-AB sharply decreased the accumulation of NF-κB p65 in nucleus (P<0.05). In addition,3-AB significantly decreased the mRNA and protein expressions of IL-1β and COX-2 which obviously increased in hippocampus at 6 h after epilepsy seizures (P<0.05). CONCLUSION:Seizures triggers NF-κB nucleus translocation and promotes the expressions of IL-1β and COX-2 in hippocampus. In addition,poly (adenosine diphosphate-ribose) polymerase inhibition by 3-AB suppresses NF-κB associated inflammatory pathway in epileptic rat hippocampus.
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AIM:To investigate the time course of nuclear factor-?B (NF-?B) and the effects of 3-aminobenzamide (3-AB) on the expressions of NF-?B,interleukin-1? (IL-1?) and cyclooxygenase-2 (COX-2) in hippocampus after seizures. METHODS:Epilepsy were induced by kainic acid through cerebral ventricular injection. Western blotting was used to detect NF-?B p65 expression in nucleus at various experiment groups. Moreover,mRNA and protein expressions of IL-1? and COX-2 in different experiment groups were determined by RT-PCR and Western blotting analysis. RESULTS:NF-?B p65 immunoreactivity began to increase in the nuclear fraction at 2 h (P