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Primary pulmonary lymphoepithelioma-like carcinoma (PPLELC) is a distinct type of lung cancer with histological profiles similar to nasopharyngeal carcinoma. The development is associated with EBV and plasma EBV DNA has predictive value in the progression and prognosis of PPLELC. PPLELC is different from some other types of lung cancer in that it has a low mutation rate of the classical lung cancer driver genes and targeted therapy is ineffective for it. Chemotherapy combined with immunotherapy may be the best first-line treatment option for patients with advanced PPLELC.
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Objective:To evaluate the inhibitory effect of a retinoid derivative ECPIRM on proliferation of a cutaneous T-cell lymphoma (CTCL) cell line HH, and to explore its potential mechanisms.Methods:Cultured HH cells were treated with ECPIRM at different concentrations of 0 (control group) , 5, 10 and 20 μmol/L separately for 72 hours, cell counting kit-8 (CCK8) assay was performed to evaluate the effect of ECPIRM on the proliferative activity of HH cells, and flow cytometry to investigate the effect of ECPIRM on apoptosis of HH cells. Some HH cells were treated with 10 μmol/L ECPIRM for 72 hours, transcriptome sequencing was performed to investigate gene expression changes triggered by ECPIRM in HH cells, and Kyoto encyclopedia of genes and genomes (KEGG) pathway analysis and gene ontology (GO) enrichment analysis were then performed to analyze differentially expressed genes in HH cells induced by ECPIRM. Reverse transcription-qPCR was subsequently conducted to verify changes in key gene expression in related pathways. Intergroup differences were analyzed by using one-way analysis of variance, and least significant difference (LSD) - t test was used for multiple comparisons. Results:CCK8 assay showed that the 50% inhibitory concentration (IC50) of ECPIRM on HH cells was 4.91 ± 2.48 μmol/L, the viability of HH cells significantly differed among the control group, and 5-, 10-and 20-μmol/L ECPIRM groups (100.00% ± 2.87%, 49.58% ± 4.53%, 48.36% ± 2.88%, 31.44% ± 2.46%, respectively, F=162.86, P < 0.001) , and was significantly lower in the 5-, 10-and 20-μmol/L ECPIRM groups than in the control group ( t=15.36, 15.73, 20.89, respectively, all P < 0.001) . Flow cytometry showed that there was a significant difference in the apoptosis rate among the 4 groups (11.51% ± 1.84%, 23.83% ± 5.72%, 36.19% ± 8.33%, 49.75% ± 4.10%, respectively, F=17.62, P < 0.001) , and the 10-and 20-μmol/L groups showed significantly increased apoptosis rates compared with the control group ( t=4.46, 6.92 respectively, both P < 0.01) . Transcriptomics analysis revealed that the inhibitory effect of ECPIRM on the cellular proliferative activity may be related to the metabolic regulation of steroids. As reverse transcription-qPCR revealed, the 10-μmol/L ECPIRM group showed significantly decreased mRNA expression of L-amino acid oxidase (IL4I1) , acetyl-coenzyme A acetyltransferase 2 (ACAT2) , 3-hydroxy-3-methylglutaryl-coenzyme A synthase 1 (HMGCS1) , mevalonate diphosphate decarboxylase (MVD) , 3-β-hydroxysteroid-8,7-isomerase (EBP) , very low-density lipoprotein receptor (VLDLR) , 3-hydroxy 3-methylglutaryl-CoA reductase (HMGCR) compared with the control group (all P < 0.05) . Conclusion:The retinoid derivative ECPIRM exerted marked anti-proliferative and apoptosis-inducing effects on HH cells, which may be related to the decreased expression of key genes involved in steroid metabolism.
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Organoselenium compounds are bioactive substances with extensive physiological activities .As a representa-tive compound ,ebselen could be used as mimics of glutathione peroxidase ,and in the treatment of many diseases ,such as car-diovascular and cerebrovascular diseases ,inflammation and noise-induced hearing loss .the research progress in physiological activities and synthetic methods of ebselen were reviewed in this paper .
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Objective To estimate the effect of a tretinoin derivative ECPIRM on retinoic acid receptors (RARs),and to observe skin irritation responses to it in mice.Methods Cultured SCL-1 cells were divided into 2 groups to be treated with culture medium containing 10 μmol/L ECPIRM (ECPIRM group) or 10 μmol/L all-trans retinoic acid (ATRA) (ATRA group) for 24 hours,and those treated with drug-free culture medium served as the control group.Western blot analysis and real-time fluorescence-based quantitative PCR were performed to quantify the protein and mRNA expressions of RARs (RARα,RARβ,RARγ and RXRα) respectively.In addition,real-time fluorescence-based quantitative PCR was conducted to measure the mRNA expressions of two target genes of the activated RAR signaling pathway,i.e.,cytochrome P450 26A1 (CYP26A1) and tazarotene-induced gene 1 (TIG1).Eight BALB/c mice were equally divided into 2 groups to be topically treated with 0.075％ ECPIRM gel or 0.05％ ATRA cream at equal molar concentrations on the shaved skin once daily for 21 successive days.Skin irritation reactions were assessed in these mice.Results Compared with the control group,the ATRA group showed significantly increased protein and mRNA expressions of RARα,RARβ and RARγ (all P ＜ 0.05).The mRNA expressions of CYP26A1 and TIG1 genes in the ATRA group were 25.49 and 3.88 times that in the control group respectively (both P ＜ 0.01).However,there was no significant difference in the protein expressions of RARα,RARβ,RARγ and RXRα,or mRNA expressions of RARα,RARβ,RARγ CYP26A1 and TIG1 between the ECPIRM group and control group (all P ＞ 0.05).Obvious Skin irritation reactions such as erythema and desquamation were observed in BALB/c mice after 2-day topical treatment with ATRA cream,and their degree peaked after 5-day treatment.However,neither erythema nor desquamation was observed in BALB/c mice during 21-day treatment with 0.075％ ECPIRM gel.Conclusion Unlike ATRA,ECPIRM cannot activate the canonical RAR signaling pathway or cause skin irritation reactions.
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Objective To investigate the effect of adenovirus-mediated IL-24 (Ad-IL-24)gene expression on the apoptosis in a human squamous cell carcinoma cell line COLO 16, and to explore the underlying molecular mechanism. Methods Cultured COLO 16 cells were divided into two groups to be transfected with an adenovirus vector carrying the IL-24 gene (Ad-IL-24 group)or green fluorescent protein (Ad-GFP group), while those receiving no treatment served as the control group. After culture for different durations, qPCR was performed to quantify IL-24 gene expression, methyl thiazolyl tetrazolium (MTT) assay to evaluate the proliferative activity of COLO 16 cells, flow cytometry to detect the apoptosis of COLO 16 cells, laser scanning confocal microscopy (LSCM) to observe the morphological changes of COLO 16 cells, Western blot to determine the levels of Bax and Bcl-2 proteins and to evaluate the activation of caspase-3, qPCR to determine the levels of Bax and Bcl-2 mRNAs, an immunofluorescence assay to observe the expression of Bax and Bcl-2 proteins. Statistical analysis was carried out by a two-sample t-test with the SPSS 19.0 software. Results MTT assay showed that the proliferation of COLO 16 cells in the Ad-IL-24 group was significantly inhibited as early as 4 days after the transfection; thereafter, the inhibitory effect increased in a time-dependent manner, and peaked on day 6(P0.05). Flow cytometry revealed that the apoptosis rate was significantly higher in the Ad-IL-24 group(13.10%± 0.92%)than in the control group(3.69%± 0.36%, P0.05). LSCM demonstrated that the apoptosis of COLO 16 cells was accelerated in the Ad-IL-24 group. The immunofluorescence assay, Western blot and qPCR all showed that the mRNA and protein expressions of Bax were increased, but those of Bcl-2 were decreased in the Ad-IL-24 group compared with the Ad-GFP group and control group. Moreover, Western blot showed a protein band that could specifically bind to the anti-cleaved caspase-3 antibody in the Ad-IL-24 group, but not in the Ad-GFP group or control group. Conclusions Ad-IL-24 can induce apoptosis in human COLO 16 squamous cell carcinoma cells, probably by up-regulating Bax expression, down-regulating Bcl-2 expression, and activating caspase 3.
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Objective To study the effect of paeonol on the proliferation and apoptosis of A375 human melanoma cells and its mechanism.Methods Cell counting kit-8 (CCK8) was used to evaluate the proliferative activity of A375 cells treated with paeonol of 0.5,1,2,4,8 mmol/L for 24,48 and 72 hours respectively.Subsequently,A375 cells were treated with paeonol of 1.25,2.5 and 5 mmol/L for 24 hours followed by double staining with annexin V and propidium iodide for the detection of cell apoptosis,fluorometric assay for the estimation of caspase 3,caspase 8 and caspase 9 activity,and Western blot for the determination of the levels of p53,nuclear factor-κB proteins and some of their target proteins.The A375 cells receiving no treatment served as the blank control group.Statistical analysis was carried out by t test.Results Within the investigated concentration and time ranges,paeonol significantly inhibited the proliferative activity of A375 cells in a concentration-and timedependent manner.Compared with the blank control group,a significant increase was observed in the early apoptosis rate in A375 cells treated with paeonol of 1.25,2.5 and 5 mmol/L for 24 hours (13.74％-± 1.73％,25.95％ ± 0.57％ and 46.44％ ± 0.81％ vs.3.11％ ± 0.53％,P ＜ 0.05 or 0.01),as well as in the activity of caspase 3,8 and 9 in A375 cells treated with paeonol of 2.5 and 5 mmol/L for 24 hours (P ＜ 0.05 or 0.01).After 24-hour treatment,the protein levels of p53 and Bax were elevated,but those of nuclear factor-κB,Bcl-2 and Bcl-XL were decreased in A375 cells with the increase of paeonol concentration.Conclusions Paeonol can inhibit the proliferation but induce the apoptosis of A375 cells,and the apoptosis-inducing effect may be realized through intrinsic and extrinsic pathways by modulating nuclear factor-κB and p53 genes.
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Objective To study the effects of triptolide on the apoptosis in and proliferation of a human melanoma cell line M14.Methods M14 cells were cultured with the presence of 5 concentrations (12.5,25,50,100,200 nmol/L) of triptolide for 24,48 and 72 hours respectively,and cell counting kit-8 (CCK-8) was used for the detection of cell proliferation.Some M14 cells were treated with triptolide at 10 nmol/L,20 nmol/L and 30 nmol/L for 48 hours followed by the analysis of cell cycle by flow cytometry and detection of cell apoptosis by flow cytometry following annexin V-fluorescein isothiocyanate (FITC)/propidium iodide double staining.The morphological changes of M14 cells treated by triptolide at 30 nmol/L for 48 hours were observed by Hoechest 33258 staining.Results Compared with untreated M14 cells,an increase of cell population in S phase was observed in triptolide-treated cells,along with a decline in cell population in G2/M phase.The apoptosis rate was (2.92 ± 0.17)％,(20.99 ± 0.40)％,(34.28 ± 2.04)％ and (63.38 ± 0.71) ％ respectively in M14 cells treated with triptolide at 0,10,20 and 30 nmol/L for 48 hours,suggesting that triptolide enhanced the proliferation of M14 cells in a dose-dependent manner.After treatment with triptolide of 30 nmol/L,M14 cells showed morphological changes characteristic of apoptosis.Conclusion Triptolide could inhibit the proliferation of and induce the apoptosis in M14 human melanoma cells.
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Aim: A new derivative LC-MS method was developed for the simultaneous determination of zofenopril and its active metabolite zofenoprilat to investigate the pharmacokinetic characteristics of zofenopril and zofenopri-lat in healthy Chinese volunteers after single and multiple oral doses of zofenopril calcium tablets. Methods: Ten Chinese healthy volunteers were given three single oral doses of 15,30, and 60 mg, respectively, and consecutively the multiple doses of 30 mg. The concentration and pharmacokinetic parameters of both the parent drug and the active metabolite were simultaneously determined by derivative LC-MS method using p-bromophenacyl bromide (p-BPB) as the derivative reagent. Results: After the single oral administrations of 15, 30, and 60 mg of zofeno-pril calcium, there was no significant difference in the t_(1/2) of both zofenopril and zofenoprilat among the three do-ses. The values of AUC_(0-24h) and c_(max) for both zofenopril and zofenoprilat showed the good linearities to the dosage over the dose range from 15 mg to 60 mg. There were no significant differences in AUC_(0-24h) and c_(max) for both com-pounds between female and male volunteers. After multiple oral administration( 30 mg once daily for 6 days ), the average steady state plasma concentration( c_(av)) for zofenopril was (5. 07 ±1. 06) ng/mL with the degree of fluctu-ation (DF) of 14. 26 ± 2. 94. The c_(av) for zofenoprilat was (6. 28 ± 1. 87) ng/mL with the DF of 11. 61 ±4. 68. The accumulation index values for zofenopril and zofenoprilat were 0. 94 ± 0. 31 and 0. 83±0. 13, respec-tively. Conclusion: Both zofenopril and zofenoprilat were demonstrated of linear kinetics after single administra-tion and showed no accumulation after multiple administration of the test zofenopril calcium tablets. There was significant difference in the pharmacokinetic characteristics for zofenopril calcium between healthy Chinese and European volunteers.
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A highly sensitive, rapid and selective liquid chromatography-tandem mass spectrometry(LC-MS/MS) method for the quantitative determination of retinamido-ester in rat plasma was developed and validated. A simplified protein precipitation with acetonitrile was employed for the sample preparation.The separation was carried out on an Agilent TC C18 column ( 150 mm×4. 6 mm ID, 5 μm particle size)with the mobile phase consisted of methanol-water-formic acid (93 : 7 : 0.1). Simvastatin was used as internal standard. The detection was performed on a trap-quadrupele tandem mass spectrometer by selected reaction monitoring (SRM) scan mode via atmospheric pressure chemical ionization (APCI). The range of and inter-day precision values were between 95.97% and 104. 43%, and RSD was between 4. 63% and 10. 69%, respectively. This method was applied to determine the pharmacokinetic parameters. The main pharmacokinetic parameters of retinamido-ester after oral administration via gastric gavage of 2. 5, 5, 10mg·kg-1 were as follows,T1/2;(11.28±7.23),(8.90±3.82),(8.01±5.56)h; AUC0-∞:(103.41±61.46),(190.23±74.99),(421.66±299.20)ng·h·mL-1;MRT:(6.31±0.75),(5.98±0.71), (6.18±0.97) h; CL/F: (30. 10 ± 13.67), (29.58±10.59), (31.18 ±17.51)L·h-1·kg-1;Vd/F:(414.94±159.82),(356.16±139.85),(369.28±322.73)L·kg-1,respectively.
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Objective To study the effects of paeonol on the tyrosinase activity and melanogenesis in co-culture model of human melanocytes and keratinocytes. Methods Melanocytes and the co-culture model of human melanocytes and keratinocytes were cultivated and the proliferation of melanocytes and the co-cultures was measured by MTT colorimetric assay. The tyrosinase activity and melanin level were measured by enzymic method. Results The melanin synthesis and tyrosinase activity were markedly suppressed by paeonol in a dose-dependent manner at the concentrations of 50?mol/L, 100?mol/L, and 200?mol/L in both melanocytes and co-cultures. The significant stronger suppression was observed with 100?mol/L and 200?mol/L of paeonol than that with controls (P
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Objective To investigate the skin reactions induced by to pical 0.1% tretinoin and its vehicle in mice.Methods 0.1% tretinoin and its v ehicle were applied separately on mice backs,once a day for 30 days.The skin r eactions,including erythema and scaling,were observed macroscopically and meas ured by biological instruments.Results Macroscopically the skin reactions of erythema and scaling induced by topical tretinoin occurred on day 3 after applic ation,reached its peak on day 6 and disappeared on day 12,and neither erythema nor abnormal scaling appeared in spite of continuing application until day 30.The erythema value measured was enhanced significantly on day 1 after applicatio n,reached its peak on day 6,maintained until day 9 and became normal on day 11.From then to day 30,it maintained at the normal level.The scaling value measu red enhanced significantly on day 3 after application,reached its peak on day 9 and became normal on day 27.Topical application of the vehicle induced neither skin reaction nor any change of erythema and scaling value.Conclusions Topic al application of 0.1% tretinoin induces skin reaction characterized by erythema and scaling.The reaction,transient but not related to the vehicle,experience s a process of "occurrence-peak-disappearance".
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Objective To evaluate the inhibitory action of ethanolic extracts from196kinds of Chi-nese herbs on tyrosinase.Methods Tyrosinase inhibitory activity was determined by the dopachrome method using L-DOPA as the substrate and the amount of dopachrome in the reaction mixture was measured by spec-trophotometer.Results Nine Chinese crude extracts[Glycyrrhiza glabra L.,Melaphis chinensis(Bell.)Baker,Cryptotympana atrata Fabricius,Paeonia suffruticosa Andr.,Sophora flavescens Ait.,Spirea thunbegii Sieb.et Bl.,Xanthium sibiricum Patr.,Morus alba L.,Rheum palmatum L.]showed potent inhibitory action on tyrosi-nase compared with positive control arbutin(1mmol/L,P0.05).Conclusion The results suggest that20kinds of crude extracts from Chinese herbs[Glycyrrhiza glabra L.,Melaphis chinensis(Bell.)Baker,Cryptotympana a-trata Fabricius,Paeonia suffruticosa Andr.,Sophora flavescens Ait.,Spirea thunbegii Sieb.et Bl.,Xanthium sibiricum Patr.,Morus alba L.,Rheum palmatum L.,Artemisia anomala S.Moore,Hemistepta lyrata Bunge,Lycium chinensis Mill.,Dipsacus asper wall.,Gastrodia elata Bl.,Forsythia suspensa(Thunb.)Vahl.,Acer gin-nala Maxim.,Glycyrrhiza uralensis Fisch,Patrinia sabiosaefolia Fisch.,Buxus sinica(Rehd.et Wils.)Ching,Lonicera japonica Thunb.,]inhibit tyrosinase and may be used as depigmentaty agents for pigmentary skin dis-eases caused by abnormal tyrosinase activity.