Effects of deguelin on the proliferation and apoptosis of ovarian cancer SKOV3 cells by regulating miR-520a-3p / 中国肿瘤生物治疗杂志

@#[摘 要] 目的：探讨鱼藤素通过调控miR-520a-3p表达对卵巢癌SKOV3细胞增殖和凋亡的影响。方法：将SKOV3细胞分为对照组（鱼藤素0 μmol/L）、鱼藤素低剂量（5 μmol/L）、中剂量（10 μmol/L）、高剂量（20 μmol/L）组，miR-NC组、过表达miR-520a-3p组，鱼藤素+anti-miR-NC组、鱼藤素+anti-miR-520a-3p组。CCK-8法、细胞集落形成实验、FCM以及qPCR法分别检测SKOV3细胞的增殖抑制率、细胞克隆形成数、凋亡率以及miR-520a-3p表达水平。结果：与对照组比较，鱼藤素（低、中、高剂量）组SKOV3细胞增殖抑制率、凋亡率、miR-520a-3p表达水平均显著升高（均P<0.05），细胞克隆形成数显著减少（P<0.05）。与miR-NC组比较，过表达miR-520a-3p组SKOV3细胞的增殖抑制率、凋亡率均显著升高（均P<0.05），细胞克隆形成数显著减少（P<0.05）。与鱼藤素+anti-miR-NC组比较，鱼藤素+anti-miR-520a-3p组SKOV3细胞的增殖抑制率、凋亡率均显著降低（均P<0.05），细胞克隆形成数显著增多（P<0.05）。结论：鱼藤素通过增加miR-520a-3p表达抑制卵巢癌SKOV3细胞的增殖能力，并诱导其凋亡。

LncRNA ZFAS1 contributes to osteosarcoma progression via miR-520b and miR-520e-mediated inhibition of RHOC signaling

ABSTRACT Objectives: We examined the expression of Lnc-ZFAS1 in osteosarcoma and comprehensively evaluated its effects on osteosarcoma in vitro and vivo. Moreover, we revealed the regulatory mechanism between Lnc-ZFAS1 and miR-520b/miR-520e-mediated RHOC and provided a novel clue for ameliorating osteosarcoma. Method: The expression of Long non-coding RNA Zinc Finger Antisense 1 (LncRNA ZFAS1) osteosarcoma tissues and normal tissues in the TCGA database was analyzed. Then, LncRNA ZFAS1 expression was further verified in clinical samples and osteosarcoma cell lines (U2OS and KHOS), as well as the human osteoblast cell line hFOB1.19 by qRT-PCR. Thereafter, LncRNA ZFAS1 was overexpressed or silenced to explore its effects on cell proliferation, apoptosis, migration, invasion, and Epithelial-Mesenchymal Transition (EMT). The fundamental mechanism through which Lnc-ZFAS1 affects osteosarcoma progression was further investigated and verified. Results: We found that LncRNA ZFAS1 was upregulated in osteosarcoma, and Lnc-ZFAS1 overexpression facilitated osteosarcoma cells proliferation, migration, invasion and EMT, while Lnc-ZFAS1 silence exerted reverse influence. Mechanistically, Lnc-ZFAS1 functionally acted as a sponger of microRNA-520b (miR-520b) and micro-RNA-520e (miR-520e) to up-regulate Ras Homologue C (RHOC). In addition, depleted Lnc-ZFAS1 restrained osteosarcoma cells proliferation, migration, and invasion, which could be rescued by RHOC overexpression. Lnc-ZFAS1 was upregulated in osteosarcoma and Lnc-ZFAS1 could exert promoted impact upon osteosarcoma cells proliferation, migration, invasion, and EMT in vitro. Conclusions: Lnc-ZFAS1 acted sponger of miR-520b and miR-520e to promote RHOC, indicating that Lnc-ZFAS1/miR-520b/RHOC and Lnc-ZFAS1/miR-520e/RHOC axes might serve as potential therapeutic strategies against osteosarcoma.

Piperine treating sciatica through regulating inflammation and MiR-520a/P65 pathway / 中国天然药物

Although the etiology of sciatica remains uncertain, there is increasing evidence that the disease process of sciatica is associated with the levels of inflammatory factors. Piperine, an alkaloid isolated from Piper nigrum, has previously been demonstrated to inhibit inflammation and analgesic effects. The purpose of this study is to verify the regulatory relationship between miR-520a and p65 and to explore how miR-520a/P65 affects the level of cytokines under the action of piperine, so as to play a therapeutic role in sciatica. Through ELISA experiment, we confirmed that four inflammatory factors (IL-1β, TNF-α, IL-10, TGF-β1) can be used as evaluation indexes of sciatica. The differentially expressed miRNA was screened as miR-520a, by microarray technology, and the downstream target of miR-520a was P65 by bioinformatics. Real-time fluorescence quantitative PCR confirmed that the expression of miR-520a was negatively correlated with pro-inflammatory cytokines, positively correlated with anti-inflammatory cytokines and negatively correlated with p65 expression at mRNA level. The expression of p65 was positively correlated with pro-inflammatory cytokines and negatively correlated with anti-inflammatory cytokines at the protein level verified by ELISA and Western blot. HE staining analysis showed that the nerve fibers were repaired by piprine, the vacuoles were significantly reduced, and the degree of nerve fiber damage was also improved. Immunohistochemical analysis showed that the expression of p65 decreased after administration of piperine. Dual-luciferase reporter gene assay confirmed that the luciferase signal decreased significantly after cotransfection of miR-520a mimics and p65 3'UTR recombinant plasmid. To sum up, in the rat model of non-compressed lumbar disc herniation, piperine plays a significant role in analgesia. MiR-520a can specifically and directly target P65, and piperine can promote the expression of miR-520a, then inhibit the expression of p65, down-regulate the pro-inflammatory factors IL-1β and TNF-α, and up-regulate the effects of anti-inflammatory factors IL-10 and TGF-β1, so as to treat sciatica.

Expression of lncRNA LOC440173 in non-small cell lung cancer tissues and its influence on the maligant biological behaviors of cancer cells / 中国肿瘤生物治疗杂志

@#[摘 要] 目的：检测lncRNA LOC440173在NSCLC组织和细胞中的表达及探讨其对癌细胞恶性生物学行为的影响。方法：选取河北医科大学第四医院生物标本库中2014至2017年手术切除的72例NSCLC患者的癌及癌旁组织标本，应用qPCR法检测NSCLC组织和癌旁组织中，以及6种NSCLC细胞株（H520、H358、A549、HCC827、H1703和H1299）中LOC440173的表达水平；构建LOC440173的敲低及过表达载体，分别转染H520和H1703细胞，应用MTS、克隆形成及Transwell小室迁移和侵袭实验分别检测敲低及过表达LOC440173对NSCLC细胞增殖、迁移及侵袭能力的影响，qPCR法检测LOC440173对于EMT过程相关标志物（E-cadherin、N-cadherin及vimentin）mRNA表达水平的影响，WB法检测其对E-cadherin、N-cadherin蛋白表达的影响。结果：LOC440173在NSCLC组织中的表达明显高于癌旁组织（P<0.01），并与淋巴结转移、组织学分化程度、TNM分期和肿瘤大小有关联（P<0.05或P<0.01）。敲低LOC440173可以抑制H520细胞的体外增殖、迁移和侵袭（P<0.05或P<0.01），过表达LOC440173可显著促进H1703细胞的增殖、迁移和侵袭（P<0.05或P<0.01）。在转录水平上，敲低LOC440173后，E-cadherin的表达水平升高，间充质相关标志物N-cadherin、vimentin的表达水平降低（P<0.05或P<0.01）；而过表达LOC440173后，E-cadherin的表达水平降低，间充质相关标志物N-cadherin、vimentin的表达水平升高（P<0.05或P<0.01）。在转录后水平上，LOC440173负向调节E-cadherin蛋白的表达、正向调节N-cadherin的蛋白表达（均P<0.05）。结论：LOC440173在NSCLC组织中的异常高表达可能与NSCLC的发生发展有关，LOC440173可显著提高NCSCL细胞的体外增殖、迁移、侵袭能力，且其作用机制可能与调控EMT相关基因表达有关。

lncRNA HOXA-AS2 regulates proliferation, migration and invasion of ovarian cancer cells via targeting miR-520a-3p / 中国肿瘤生物治疗杂志

@#[Abstract] Objective:To explore the targeting relationship between long-chain noncoding RNA HOXA-AS2 (lncRNA HOXA-AS2) and microRNA-520a-3p (miR-520a-3p) and their effects on the proliferation, migration and invasion of ovarian cancer SKOV3 cells. Methods： ：qPCR was used to detect the expression levels of lncRNA HOXA-AS2 and miR-520a-3p in various ovarian cancer cell lines (SKOV3, HO8910, OVCAR3 cells) and normal ovarian epithelial cell line HOSE. Bioinformatics methods were used to predict the targeting relationship between HOXA-AS2 and miR-520a-3p, which was then verified by Dual luciferase reporter gene assay. si-HOXA-AS2, miR-520a-3p mimic, anti-miR-520a-3p and corresponding control fragments were transfected into SKOV3 cells separately or in combination. MTT, Transwell and Western blotting were used to detect the proliferation, migration, invasion and expressions of related proteins (CyclinD1, p21, p27, MMP-2, MMP-9, MMP-14) of SKOV3 cells in each group. Results: Compared with HOSE cells, HOXA-AS2 was over-expressed while miR-520a-3p was under-expressed in ovarian cancer cell lines (all P<0.05). HOXA-AS2 could targetedly down-regulate the expression of miR-520a-3p. Compared with the NC group, the proliferation, migration and invasion of SKOV3 cells in the si-HOXA-AS2 and miR-520a-3p mimics groups were significantly reduced (all P<0.01), and the protein expressions of p21 and p27 were significantly increased, while protein expressions of CyclinD1, MMP-2, MMP-9, MMP-14 were significantly reduced (all P<0.01). The proliferation, migration and invasion of SKOV3 cells in the si-HOXA-AS2+antimiR-520a-3p group were significantly enhanced compared with those in si-HOXA-AS2 and si-HOXA-AS2+anti-miR-NC groups (all P<0.05). Conclusion: lncRNA HOXA-AS2 enhances the proliferation, migration and invasion of ovarian cancer SKOV3 cells by targetedly inhibiting the expression of miR-520a-3p.

Cinobufagin suppresses metastasis and invasion of esophageal carcinoma Kyse-520 cells through FAK/PI3K/AKT pathway / 中国药理学通报

Aim To investigate the effect of cinobufagin on the migration and invasion of esophageal cancer Kyse-520 cells, and to explore the underlying molecular mechanism. Methods The rates of inhibition after treated with different concentrations of cinobufagin 12, 24, 48 h were detected by CCK-8 method, and the changes of cell migration and invasion were observed with wound healing and transwell assay. The mRNA expressions of FAK, Akt, PTEN, VEGF-A, MMP-2 and MMP-9 were detected by quantitative real-time polymerase chain reaction ( RT-qPCR ). The protein expressions of FAK, p-FAK ( Tyr397 ), Akt, p-Akt (Sei473 ), VEGF-A, PTEN, MMP-2 and MMP-9 were measured by Western blot. Results The results of CCK-8 showed that cinobufagin could inhibit the proliferation of Kyse-520 cells in a time- and concentration-dependent manner, and cinobufagin significantly inhibited the cell invasion and migration. Meanwhile, data from RT-qPCR and Western blot suggested that cinobufagin had no significant effect on mRNA and total protein of FAK and Akt, but it reduced the expression of p-FAK(Tyr397), p-Akt(Ser473), VEGF- A, MMP-2 and MMP-9, and increased the PTEN expression. Conclusions Cinobufagin significantly inhibits the invasion and migration of esophageal cancer Kyse-520 cells through inducing PTEN expression and down-regulating FAK/PI3K/AKT pathway.

miR-520a-3p enhances the paclitaxel sensitivity of non-small cell lung cancer A549/TAX cells by targeting FZD8 / 中国肿瘤生物治疗杂志

@#Objective: To investigate the influence of miR-520a-3p on paclitaxel (TAX) sensitivity of non-small cell lung cancer (NSCLC)A549/TAX cells via regulating frizzled class receptor 8 (FZD8). Methods: NSCLCA549 cells, TAX-resistant cell lineA549/ TAX and human lung epithelial HLF-α cells were selected. The expression level of miR-520a-3p in A549 and A549/TAX cells was detected by qPCR. According to different transfection plasmids, the experimental cells were divided into control group, miR-520a-3p mimics group, si-FZ8 group and si-FZD8+miR-520a-3p inhibitor group. After being treated with 6 μmol/L paclitaxel, the proliferation ofA549/TAX cells was determined by CCK-8 assay. Flow cytometry withAnnexin V-FLTC/PI staining was used to detect the apoptosis level of A549/TAX cells. The expression of FZD8 in A549/TAX cells was detected by WB. The targeting relationship between miR520a-3p and FZD8 was verified by the dual-luciferase reporter gene system. Results: miR-520a-3p was poorly expressed in TAX-resistant A549/TAX cells (P<0.01), and TAX up-regulated the expression of miR-520a-3p in A549/TAX cells (P<0.01). After the treatment with 6 μmol/L TAX, over-expression of miR-520a-3p significantly inhibited the proliferation of A549/TAX cells and promoted apoptosis (all P<0.01). Dual luciferase reporter gene assay showed that miR-520a-3p targetedly down-regulated the expression of FZD8 (P< 0.01). si-FZD8 could significantly inhibit the proliferation and promote cell apoptosis of A549 / TAX cells, thereby enhancing the TAX sensitivity of cells. At the same time, simultaneous knockdown of miR-520a-3p and FZD8 could reverse the enhancement of FZD8 knockdown on TAX sensitivity of A549/TAX cells (P<0.01). Conclusion: miR-520a-3p enhances the TAX sensitivity of A549/TAX cells by down-regulating the expression of FZD8.

miR-520d over-expression reverses chemotherapy resistance of TNBC via inhibiting autophagy protein Beclin1 / 中国肿瘤生物治疗杂志

@# Objective: To investigate the role and molecular mechanism of miR-520d in reversing the chemoresistance of triple negative breast cancer (TNBC) by regulating autophagy. Methods: Docetaxel (Doc) resistant cell lines MDA-MB-231/Doc and MDA-MB468/Doc were constructed by using human TNBC cell lines MDA-MB-231 and MDA-MB-468 as parental cells, and the cells were divided into blank group (parental cells), control group (drug-resistant group), and miR-520d over-expression group. The expression levels of miR-520d in cells of the blank and drug-resistant groups were detected by qPCR. The Doc-sensitivity of resistant cells over-expressing miR-520d was detected by MTT assay.After MDC staining, the generation of autophagosome in cells was observed under fluorescence microscopy; the number of miR-520d over-expressed resistant cells with positive LC3 expression was observed under confocal microscopy. The luciferase reporter gene assay was used to verify the targeting relationship between miR-520d and Beclin1. The effect of miR-520d mimics on the expression of autophagy-associated protein Beclin1, and LC3Ⅰ, LC3Ⅱ in cells was detected by WB assay. Results: The results of qPCR showed that the expression of miR-520d in the drug-resistant TNBC cells was significantly lower than that of normal cells (P<0.01). In drug-resistant cells over-expressing miR-520d, the Doc-sensitivity was significantly improved, while the autophagy activity was significantly reduced (all P<0.01).At the same time, luciferase experiments demonstrated that Beclin1 was a possible target molecule of miR-520d (P<0.05). WB results showed that the combination of docetaxel and miR-520d mimics reduced the LC3-II/I ratio and the expression of autophagy protein Beclin1 in drug-resistant TNBC cells (all P<0.05). Conclusion: The regulation of miR-520d levels may alter the expression of autophagy protein Beclin1, thereby reversing Doc chemotherapy resistance in TNBC cells.

Construction of lung cancer cell model overexpressing human MutS homologue 2(hMSH2) / 基础医学与临床

Objective To construct human lung cancer cell model with human MutS homologous protein 2 (hMSH2) overexpression for exploring the effect of hMSH2 molecule in the cytotoxicity of γδ T cell against lung cancer cells.Methods hMSH2 coding sequence was cloned by PCR for construction of recombinant vector which over expressed hMSH2-EGFP fusion protein using homologous recombination.The recombinant vector was transfected to lung cancer cell line NCI-H520 to construct human lung cancer cell model overexpressing hMSH2 molecule.The expression of hMSH2 molecule in NCI-H520 was detected by Western blot.The expression of hMSH2 on the cell membrane was measured by flow cytometry.Cytotoxicity of expanded γδ T cells from peripheral blood mononuclear cells against NCI-H520 cells was detected by LDH release assay in vitro.Results hMSH2 coding sequence (2805 bp) was cloned and the result of restriction endonuclease digestion of Fugw-hMSH2 recombinant vector was accordance with the anticipated objective strip size.Exogenous hMSH2-EGFP fusion protein was expressed in NCIH520 cells.The level of hMSH2 molecule on the surface of NCI-H520 cells with overexpression of hMSH2 was significantly increased (P＜0.001).Cytotoxicity of γδ T cells against NCI-H520 cells with overexpression of hMSH2 was significantly increased compared to the wild type NCI-H520 cells (P＜0.05).Conclusions Lung cancer cell model that overexpresses hMSH2 molecule is successfully constructed,hMSH2 molecule on the cell membrane is increased and the cytotoxicity of γδ T cells against lung cancer cells is enhanced.

Study on effect of FTY720 combined with gecitabine on proliferation and apoptosis of non-small cell lung cancer tumor-related cell-lines / 重庆医学

Objective To investigate the effect of FTY720 and gemcitabine on the proliferation and apoptosis of H520 and A549 cells in non-small cell lung cancer(NSCLC) cell line.Methods The interventional influence on the in vitro cultured NSCLC A549 and H520 cells was performed by selecting 0,2,4,6,8,10 μmol/L concentrations of FTY720,then the absorbance value was detected at 24,48,72 h after culture and the proliferation inhibiting effects of FTY720 on A549 and H520 were observed under the condition of different concentration of FTY720;adding single 7 μmol/L of FTY720,single 0.2 μmol/L gemcitabine and 37 μmol / L FTY720 combined with 0.2 mol/L gemcitabine into A549 and H520 cells lines,then the differences of inhibition and apoptosis after 48 h in the cells of each group were observed.Results The inhibitory effect of different concentrations of FTY270 on NSCLC A549 and H520 cell lines was statistically significant difference (P＜0.05).The proliferation inhibiting effect of FTY720 on NSCLC H520 and A549 cell lines had the correlation with the concentration and time.The apoptosis rate of FTY720 combined with gemcitabine on A549 and H520 cells was significantly higher than that of single use in these two drugs (P＜0.05).Conclusion FTY720 combined with gemcitabine can significantly inhibit the proliferation of A549 and H520 in human NSCLC,and can effectively promote the apoptosis of cancer cells,and has the higher clinical value.

Effect of co-transfection of miR-520c-3p and miR-132 on proliferation and apoptosis of hepatocellular carcinoma Huh7

Objective To investigate the effects of co-transfection of miR-520c-3p and miR-132 on proliferation and apoptosis of hepatocellular carcinoma Huh7. Methods Hepatocellular carcinoma Huh7 was cultured in vitro and lipidosome was used to transfect miR-520c-3p and miR-132, respectively or together. The effects of transfection of miR-520c-3p and miR-132 on proliferation and apoptosis of Huh7 were detected by CCK8 and Annexin V staining and flow cytometry, and the expression level of the targeted gene of over-expressed miR-520c-3p and miR-132 was determined by Western blot and realtime PCR. Results Compared with the control group, the proliferation ability of Huh7 of the single transfected and co-transfected miR-520c-3p and miR-132 decreased significantly, and the apoptosis ratio increased distinctly (P < 0.05). Besides, the effect of the co-transfection group was better than that of the single transfection group. The protein levels of GPC3 (Glypican-3) and YAP (Yes-associated protein), the target genes transfected only by miR-520c-3p and miR-132, respectively, reduced obviously (P < 0.05), which was similar with the co-infected cells, but cells transfected by miR-132 only showed a decrease of YAP. Conclusions The co-transfection of miR-520c-3p and miR-132 can target-regulate the expression of GPC3 and YAP, enhance the exhibition effect on proliferation of hepatocellular carcinoma Huh7 and induce cell apoptosis synergistically.

Effect of co-transfection of miR-520c-3p and miR-132 on proliferation and apoptosis of hepatocellular carcinoma Huh7

OBJECTIVE@#To investigate the effects of co-transfection of miR-520c-3p and miR-132 on proliferation and apoptosis of hepatocellular carcinoma Huh7.@*METHODS@#Hepatocellular carcinoma Huh7 was cultured in vitro and lipidosome was used to transfect miR-520c-3p and miR-132, respectively or together. The effects of transfection of miR-520c-3p and miR-132 on proliferation and apoptosis of Huh7 were detected by CCK8 and Annexin V staining and flow cytometry, and the expression level of the targeted gene of over-expressed miR-520c-3p and miR-132 was determined by Western blot and realtime PCR.@*RESULTS@#Compared with the control group, the proliferation ability of Huh7 of the single transfected and co-transfected miR-520c-3p and miR-132 decreased significantly, and the apoptosis ratio increased distinctly (P < 0.05). Besides, the effect of the co-transfection group was better than that of the single transfection group. The protein levels of GPC3 (Glypican-3) and YAP (Yes-associated protein), the target genes transfected only by miR-520c-3p and miR-132, respectively, reduced obviously (P < 0.05), which was similar with the co-infected cells, but cells transfected by miR-132 only showed a decrease of YAP.@*CONCLUSIONS@#The co-transfection of miR-520c-3p and miR-132 can target-regulate the expression of GPC3 and YAP, enhance the exhibition effect on proliferation of hepatocellular carcinoma Huh7 and induce cell apoptosis synergistically.

Mir-520c-3p targeted GPC3 for hepatocellular carcinoma Huh-7cell proliferation, migration and invasion ability / 国际外科学杂志

Objective To research the effect of mir-520c-3p targeted GPC3 to the hepatocellular carcinoma Huh-7 cell proliferation,migration,and the influence of the attack ability and find new theoretical basis for liver hepatocellular carcinoma clinical treatment.Methods The cells were divided into three groups:not transfection of mir-520c-3p group (cell group),negative control group (Nc group),and transfection of mir-520c-3p group (treat ment group).Then used fluorescence quantitative PCR and Western Blot to detect GPC3mRNA gene and protein expression quantity.Cell proliferation of change was detected by the EDU.Made use of Transwell to detect cell invasion and migration ability of the change.Results Fluorescence quantitative PCR results showed that Cell group,NC group and treatment group were 1.13 ± 0.23,1.28 ± 0.15 and 1.05 ± 0.19 (P ＞ 0.05),mir-520-3p could not reduce the GPC3mRNA; but Western Blot detection results showed that GPC3 protein expression level reduce significantly after transfection mir-520c-3p,Cell,NC and treatment group were 2.16 ± 0.08,1.99 ± 0.04 and 0.499 ± 0.05 (P ＜ 0.01).The EDU detection results showed that hepatocellular carcinoma Huh-7 cell proliferation ability obviously inhibited after transfection mir-520c-3p,Cell group,NC group and treatment group were (90.12 ± 1.93) ％,(91.02 ± 0.35) ％ and (77.73 ± 5.88) ％ (P ＜ 0.05),and Transwell test found that hepatocellular carcinoma Huh-7cell invasion abilities were restrained,Cell group,NC group and treatment group were 0.071 ±0.008,0.105 ±0.001 and 0.048 ± 0.002 (P ＜ 0.05),in the same the cells' migration abilities were reduced,Cell group,NC group and treatment group were 0.546 ± 0.010,0.328 ± 0.002 and 0.151 ± 0.002 (P ＜0.01).Conclusions Mir-520c-3p can target GPC3 so that affect hepatocellular carcinoma Huh-7 cell proliferation,invasion and migration abilities.

Characterization of the nover late T cell activation proteins, 519 and 520

The late T cell activation gene, 519, is expressed in antigen specific, growth factor dependent T cell lines and clones but not in T or B cell tumors, other hematopoietic cells, tonsil, muscle, lung, or liver. Resting peripheral blood lymphocytes (PBLs) express little or no 519mRNA, but levels increase dramatically 5~7 days after activation with alloantigen or mitogen. Four alternatively spliced transcripts of the gene exist. 520 cDNA, the predominate gene transcript, encodes a protein of 144 amino acids and has a potentially cleavable hydrophobic leader of 15 amino acids at the amino terminus. The 519 transcript encodes a 129 amino acid product. An polyclonal antiserum was produced by immunizing a rabbit with a synthetic peptide corresponding to the second hydrophobic region common to both 519 and 520. 14.3 and 15.5kD protein bands were immunoprecipitated (IP) from radiolabled in-vitro translated products, 519 and 520 respectively. A protein of 14.3kD was detected by IP of in-vivo 35S cysteine and methionine radiolabled cytotoxic T lymphocytes (CTL) cells. Western blotting of lysates from a CTI line and a B cell tumor line revealed a distinct band at 14.3kD only in CTL lanes. Both CTL and peripheral blood lymphocytes expressed the greatest amount of 519/520 protein in cells 5~7 days after stimulation. Western blotting of differential centrifugation samples localized 519/520 protein to the granule layer. Comparison of 519/520 proteins with other known protin sequences identified homology with surfactant associated and sphyngolipid activating proteins. A homologous gene(NKC5) is present in natural killer cells (NK). The increase in the amount of the 519/520 protein product in CTL and PBL parallel increases in mRNA expression. 519/520 and related gene products are present in CTL, PBL and NK cells, have structural similarity with lipid associating proteins.