Pyruvate and cilostazol protect cultured rat cortical pericytes against tissue plasminogen activator (tPA)-induced cell death

Kim, H.N.,Kim, T.Y.,Yoon, Y.H.,Koh, J.Y. Elsevier/North Holland 2015 Brain Research Vol.1628 No.2

Since even a brief ischemia can cause permanent brain damage, rapid restoration of blood flow is critical to limiting damage. Although intravenous tPA during the acute stage is the treatment of choice for achieving reperfusion, this treatment is sometimes associated with brain hemorrhage. Agents that reduce tPA-related bleeding risk may help expand its therapeutic window. This study assessed whether zinc dyshomeostasis underlies the toxic effect of tPA on brain vascular pericytes; whether pyruvate, an inhibitor of zinc toxicity, protects pericytes against tPA-induced cell death; and whether cilostazol, which protects pericytes against tPA-induced cell death, affects zinc dyshomeostasis associated with tPA toxicity. Cultured pericytes from newborn rat brains were treated with 10-200μg/ml tPA for 24h, inducing cell death in a concentration-dependent manner. tPA-induced cell death was preceded by increases in intracellular free zinc levels, and was substantially attenuated by plasminogen activator inhibitor-1 (PAI-1) or TPEN. Pyruvate completely blocked direct zinc toxicity and tPA-induced pericyte cell death. Both cAMP and cilostazol, a PDE3 inhibitor that attenuates tPA-induced pericyte cell death in vitro and tPA-induced brain hemorrhage in vivo, reduced zinc- and tPA-induced pericyte cell death, suggesting that zinc dyshomeostasis may be targeted by cilostazol in tPA toxicity. These findings show that tPA-induced pericyte cell death may involve zinc dyshomeostasis, and that pyruvate and cilostazol attenuate tPA-induced cell death by reducing the toxic cascade triggered by zinc dyshomeostasis. Since pyruvate is an endogenous metabolite and cilostazol is an FDA-approved drug, in vivo testing of both as protectors against tPA-induced brain hemorrhage may be warranted. This article is part of a Special Issue entitled SI: Neuroprotection.

삼출성 늑막액에서 양악성 감별지표로서 CEA, TPA, SCC Ag 측정의 의의

김경찬,김민수,김미정,권두영,한승범,전영준 啓明大學校 醫科大學 1998 계명의대학술지 Vol.17 No.4

상피 세포에서 기원한 대표적인 종양표지자인 carcinoembryonic antigen (이하 CEA로 약함)은 삼출성 늑막액 환자의 양악성 감별에 보고적으로 사용되고 있다. CEA 이외에 혈청에서 양악성감별의 보조적 지표로 알려진 tissue polypeptide antigen (이하 TPA로 약함)과 squamous cell carcinoma antigen (이하 SCC Ag으로 약함)을 혈청과 늑막에서 동시 측정하여 늑막삼출액의 악성 감별에 어느 정도의 임상적인 유용성이 있는지를 알아보기 위하여 이 연구를 시행하였다. 1997년 1월 1일부터 동년 8월 31일까지 계명대학교 동산의료원에 입원한 환자들 중 삼출성늑막액을 가진 61명을 대상으로 하여 혈청과 늑막액에서 CEA, TPA, SCC Ag의 수치를 방사면역법으로 측정하였다. 각각의 조양표지자들은 악성과 양성군으로 구분한 뒤 분석하였으며 악성군이 28례, 양성군이 33례이었다. 그리고 진단양성기준치를 설정한 뒤 종양표지자들의 특이도, 민감도를 산출하였고 상기 지표들을 종양표지자와 늑막액 세포검사르 조합한 경우에도 산출하여 비교 분석하였다. 혈청 CEA 와 TPA는 각각 7.0 ng/ml, 80.0 ng/ml, 늑막액 CEA와 TPA는 각각 50. ng/ml, 4700.0 ng/ml로 진단양성기준치를 설정하였을 때 특이도를 낮추지 않으면서 가장 높은 민감도를 보였다. 늑막액 세포검사와 동시에 혈청 TPA 도는 늑막액 CEA를 측정하였을 때 특이도는 떨어뜨리지 않으면서 민감도를 높이는 좋은 조합인 것으로 나타났으며 혈청 CEA 및 TPA수치를 늑막액 세포검사와 도시에 시행하였을 때 특이도를 떨어뜨리지 않으면서 가장 높은 민감도를 얻었다. 늑막액 세포검사가 음성인 경우에도 혈청 CEA와 TPA를 동시에 측정하여 높은 민감도와 특이도를 얻을 수 있었다. 따라서 CEA와 TPA는 늑막삼출액의 양악성 감별 진단 유용한 보조적 지표로서 사용할 수 있을 것으로 판단된다. Carcinoembryonic antigen(CEA), the most widely used tumor marker was measured in pleural fluid of patients with exudative pleural effusions in order to differentiate malignant from benign effusions. This study was performed to find out if there is any clinical utility in differential diagnosis of malignancy by measuring simultaneously CEA, tissue polypeptide antigen(TPA) and squamous cell carcinoma antigen(SCC Ag) in serum and pleural fluid. The study population was 61 patients with exudative pleural effusions who were admitted to Keimyung University Hospital from January 1 to August 31, 1997. Each CEA, TPA and SCC Ag level in serum and pleural fluid were measured using radioimmunoassay method. These patients were divided to malignant and benign group. Malignant group consists of 28 cases and benign group consists of 33 cases. And the sensitivity and specificity of each tumor marker was obtained using cut-off value and that combining tumor markers and pleural fluid cytology were also obtained and analyzed. When the cut-off value was applied to CEA and TPA in serum using 7.0 ng/ml and 80.0 ng/ml respectively, the highest sensitivity was obtained without specificity being lowered. The same result was obtained when the cut-off value was applied to CEA and TPA in pleural fluid using 5.0 ng/ml and 4700.0 ng/ml respectively. When CEA in pleural fluid or TPA in serum were measured in combining with pleural fluid cytology sensitivity was increased without decreasing specificity than measured in pleural fluid cytology alone. When CEA in serum and TPA in serum were measured in combining with pleural fluid cytology simultaneously, the highest sensitivity was produced without decreasing specificity than measured in any other combinations. In addition, when serum CEA and TPA in serum were measured in the negative group of pleural fluid cytology, high sensitivity and specificity were obtained. These data suggest that CEA and TPA can be used as useful tumor markers for the differential diagnosis of malignancy and benign condition in patients with exudative pleural effusions.

Effect of 12-O-tetradecanoylphorbol 13-acetate (TPA) on the Expression of Keratin in HaCaT Cells

안수홍(Soo-Hong Ahn),이승호(Sung-Ho Lee),김대광(Dae-Kwang Kim),김주영(Joo-Young Kim) 대한해부학회 2003 Anatomy & Cell Biology Vol.36 No.4

사람 피부에서 특정한 keratin 표지자들은 각각 정상적인 분화와 병적인 상태를 반영한다. 본 실험에서는 12-Otetradecanoylphorbol 13-acetate (TPA)에 노출된 HaCaT 세포들의 형태학적인 소견들과 함께 피부의 정상적인 분화표지자인 keratin 10 (K10), 병적인 분화표지자들인 keratin 8 (K8)과 keratin 13 (K13)의 발현양상을 밝히고자 하였다. 이를 위해 다양한 농도(0, 0.1, 1 μg/ml)의 TPA를 이 세포들에 2시간 또는 6시간동안 처리하였다. TPA에 의해 다각형의 HaCaT 세포들은 섬유모세포의 형태로 변화하였다. 핵막의 심한 접힘이 나타났고, 부착반의 수는 감소하였다. 간접면역형광법과 Northern blotting의 소견에서 TPA는 각각 단백질과 mRNA 수준에서 K10의 발현을 모두 감 소시켰지만, K8과 K13의 발현은 상당히 증가되었다. 유세포분석에서 keratin과 DNA의 양을 동시에 측정하였는데, K8과 K13의 발현이 S-G2-M기에서도 현격히 증가되었다. 결론적으로 TPA에 의해 유발된 위의 결과들은 HaCaT 세포에서의 형태학적인 변화와 keratin의 발현 사이에 밀접한 관 계가 있을 가능성을 제시하였다. 또한 피부에서 종양촉진자인 TPA가 종양개시자의 도움없이 직접적으로 악성세포를 유발 시킬 수도 있을 것으로 생각된다. In human skin, specific keratin markers reflect on normal differentiation and pathologic conditions. This experiment focused on the expressional pattern of keratin 10 (K10: normal differentiation marker), and keratin 8 & 13 (K8 & K13: pathologic differentiation marker) together with their cellular localization after treating HaCaT cells with 12-Otetradecanoylphorbol 13-acetate (TPA). The cells were treated with TPA at 0, 0.1, 1 μg/ml for 2 hours or 6 hours. Morphologic studies revealed that TPA treatment changed the shape of cells into the fibroblast-like cells with highly folded nuclear membrane and reduced number of the desmosome. The results of indirect immunofluorescent staining and Northern blotting analysis showed that TPA considerably down-regulated the expression of K10, while markedly up-regulating the expression of K8 and K13 both at protein and mRNA levels. Furthermore, by simultaneous staining for keratins and DNA content in flow cytometry, it was found that TPA increased the expression of K8 and K13 dramatically at the S-G2-M phase of the cells. In conclusion, these changes induced by TPA in HaCaT cells may indicate a close relationship between the morphologic change and the altered expression of keratin subfamilies. It also suggests that TPA known as a tumor promotor may directly induce the potentially malignant cells even without the support of tumor initiator.

12-O-Tetradecanoyl phorbol-13-acetate (TPA)-induced growth arrest is increased by silibinin by the down-regulation of cyclin B1 and cdc2 and the up-regulation of p21 expression in MDA-MB231 human breast cancer cells

Kim, S.,Lee, H.S.,Lee, S.K.,Kim, S.H.,Hur, S.M.,Kim, J.S.,Kim, J.H.,Choe, J.H.,Shin, I.,Yang, J.H.,Lee, J.E.,Nam, S.J. G. Fischer 2010 Phytomedicine Vol.17 No.14

TPA is a potent regulator of cell growth, including cell proliferation and differentiation. In this study, we determined the effect of silibinin on TPA-induced growth arrest in breast cancer cells. Silibinin increased growth arrest of the G2/M phase in a dose-dependent fashion. Silibinin decreased the basal level of cyclin B1 and cdc2 expression, which is involved in S phase and G2/M transition. In addition, TPA-induced G2/M phase arrest was increased by silibinin. Under the same conditions, TPA-induced down-regulation of cyclin B1 and cdc2 was decreased by silibinin. In contrast, TPA-induced p21 expression was further increased by silibinin. To determine the regulatory mechanism of TPA-induced growth arrest, we pretreated cells with various inhibitors, such as UO126, SB203580, and LY294002. Interestingly, TPA-induced growth arrest was significantly increased by LY294002, but not by UO126 and SB203580. In addition, TPA-induced down-regulation of cyclin B1 was inhibited by LY294002; however, the basal level of p21 was increased by TPA and TPA-induced p21 expression was further increased by LY294002. Finally, adenoviral constitutively active-Akt (Ad-CA-Akt) overexpression regulated the up-regulation of cyclin B1 and the down-regulation of p21. Therefore, we have demonstrated that silibinin has an additive effect on TPA-induced growth arrest through the PI-3-kinase/Akt-dependent pathway.

자가동맥색전을 이용한 뇌색전모형 토끼에서 Tissue Plasminogen Activator의 효과

이경진,최창락 대한신경외과학회 1994 Journal of Korean neurosurgical society Vol.23 No.9

The safety and efficacy of intravenous tissue plasminogen activator(tPA) on the condition of ruling out the significant risk were studied at 6 and 12 hours after cerebral artery embolization in rabbit model. The time selection was chosen to simulate the analogus clinical situation. The safety and effectiveness of tPA in experimental and clinical treatment of acute coronary thrombosis have been established. Tissue plasminogen activator is an endogenous fibrin-specific serine protease with the potent thrombolytic activity that has been produced recently by recombinant DNA thenology. The acute cerebral thromboembolic model was induced by injecting three 0.5×1.0㎜ fragments of autologous arterial thromi into internal carotid artery through the intra-arterial catheter. The autologous arterial thrombi was obtained from the traumatized arterial endothelium by scratching the lumen of auricular artery using modified spinal needle. The experimental group was divided into four groups : ① group Ia : saline-treated(1㎖/㎏) control group at 6 hours after embolization(n=10), ② group Ib : tPA-treated group(1㎖/㎏) at 6 hours after embolization(n=10), ③ group IIa : saline-treated control group at 12 hours after embolization(n=10), ④ group Iib : tPA-treated group at 12 hours after embolization(n=13). The experimental rabbits were sacrificed at 24 hours after injection of tPA(1㎖/㎏) or saline(1㎖/㎏) in each group. Brain was cut into 0.5㎝ thick coronal sections, which were stained with triphenyltetrazolium chloride to define the areas of infarction. The transparent plastic sheets were placed on the each section, and the total area of the brain slice and the area of infarction were measured by the plannimeter(as outlined by TTC staining). The percentage area of whole brain infarction was calculated as (the sum of infarcted are the sum of brain slice areas) × 100% for each rabbit. We also observed the pathologic findings with hematoxylin-eosin staining. We results were as follows : 1) Only 1 rabbit treated with tPA at 12 hours after occlusion exhibited the gross hemorrhage. 2) The infarcted area was limited to the basal ganglia and cortex in all groups. 3) The mean percentage area of whole brain infarction averaged 18.6±1.94% in group Ia, 6.32±1.02% in group Ib, and 20.8±3.34% in group IIa, 6.78±1.40% in group IIb. One-way ANOVA test of infarction size showed the significant differences(P<0.05) between the tPA-treated group and the saline-treated control group, but no difference between the group treated with same agent. 4) Under the study of microscope, infarcted area of saline-treated control group was more extended than that of tPA-treated group. Coagulation necrosis and degeneration of neuronal cells could be seen. But the infarcted area of tPA-treated group was smaller than that of saline-treated control group. Only collection of foamy macrophages adjacent the necrotic area could be seen in tPA-treated group. These results suggest that tPA therapy may be safe and efficacious during the interval of 6 to 12 hours after embolization.

황반하출혈 환자에서 조직형 플라스미노겐 활성제(tPA) 주입술 후 발생한 망막독성 1예

정종진,조성원 대한안과학회 2009 대한안과학회지 Vol.50 No.5

Purpose: To present the clinical feature of retinal toxicity of intravitreal tissue plasminogen activator which was used for treatment of submacular hemorrhage. Case summary: An intravitreal injection of tPA (100 μg) with C3F8 gas tamponade (0.2 cc) was given to treat the submacular hemorrhage in a patient with ARMD. The therapeutic effect was measured by visual acuity, slit lamp examination, indirect funduscopy and fluorescein angiogram. Three months after the operation, the hemorrhage was decreased but a pigmentary change was observed on the peripheral retina. After 8 months, the submacular hemorrhage completely reabsorbed but the peripheral pigmentary change had increased. Ten months later, the retinal pigmentary change was observed on the entire retina except the posterior pole. The fluorescein angiogram showed peripheral hyperfluorescene of the retina due to window defect from the pigmentary change but no leakage was detected. The electroretinogram showed reduced amplitude in the right eye. Conclusions: Intravitreal tPA injection of 25 to 100 μg with pneumatic displacement is typically used for the treatment of submacular hemorrhage. However, there is no established safety dose of tPA for use in human eyes. In the present study, 100 μg of tPA was used and retinal toxicity was noted. Establishing a safety dose of tPA to prevent dosage dependent complications is necessary. 목적: 황반하출혈 환자에서 시행한 유리체강내 조직형 플라스미노겐 활성제(tPA) 주입 후 망막독성이 확인되어 이를 보고하는 바이다. 증례요약: 연령관련황반변성 환자에서 발생한 황반하출혈의 치료로 100 μg의 tPA 및 C3F8 gas 주입을 시행한 후 시력검사, 세극등검사, 안저촬영 및 형광안저혈관조영술을 사용하여 10개월 동안 경과관찰을 하였다. 주입 세달 후 출혈은 감소하였으나 망막 주변부의 색소 변성이 관찰되었다. 8개월 후 출혈은 완전히 흡수되었으나 변성은 더 증가한 양상이었고 10개월 후 후극부를 제외한 전 망막에서 색소 변성을 관찰할 수 있었다. 10개월 후 시행한 형광안저혈관조영술에서 누출소견은 보이지 않았으며, 주변부 쪽으로 색소변성에 의한 과형광 소견을 보였고 망막전위도 검사 상 전체적인 우안의 진폭감소를 보였다. 결론: 황반하출혈시 사용하는 tPA의 용량은 25~100 μg까지 다양하나 사람 눈에서 안전하게 사용할 수 있는 용량은 아직 확인되지 않았다. 본 증례에서는 tPA 100 μg에 의해 망막독성이 발생되었다. 향후 전향적 연구를 통해 tPA의 안전한 용량의 정립 및 용량에 따른 합병증의 병발에 대한 연구가 필요할 것으로 사료된다.

Spatial changes of the maxillary molar following unilateral derotation with the precision TPA

Kim, You-Sun,Yeh, Seong-Pil,Kang, Dae-Woon,Chun, Youn-Sic,Row, Joon 대한치과교정학회 2004 대한치과교정학회지 Vol.34 No.3

본 연구의 목적은 치아의 이동양상을 관찰하기 위해 고안된 Calorific machine system (typodont simulation system)과 precision TPA를 이용하여, 근심 회전된 (mesial-in rotation) 상악 대구치를 회 전(derotation)시킨 후 해당 치아 및 반대측 고정원의 공간변화를 확인하기 위함이다. 우측 상악 제 1대구치를 고정원으로 사용하였고 좌측 상악 제 1대구치를 근심 회전된 치아로 사용하였으며, TPA에 부여한 회전각은 20˚, 40˚, 60˚ 였다. 먼저 수동형의 precision TPA를 제작한 후, TPA의 왼쪽 삽입부위(tag)를 각각 20˚, 40˚, 60˚로 구부려 실험하였다. 각 군별 실험은 동일한 조건에서 5회 반복한 후 ANOVA와 Tucky's Studentized Range(HSD) test로 검정하였다. 실험 결과, 교합면에서는 precision TPA의 구부리는 각도가 증가할수록 회전된 구치의 원심 설측 교두가 근심 설측 방향으로 움직이는 동안, 근심 협측 교두는 협측으로 더 많이 움직였고(p<0.001) 원심방향으로는 더 적게 움직였다.(p<0.001) 시상면에서 회전 구치의 구개측 치근은 더욱 근심으로 움직였다.(p<0.001) 횡단면에서는 회전된 치아가 약간의 정출을 보였다(p<0.001) 고정원으로 사용된 우측 상악제1대구치는 세 평면에서 의미 있는 변화를 보이지 않았다. The purpose of this study was to evaluate the spatial changes of mesial-in rotated maxillary molar and opposite anchor tooth during derotation by the precision transpalatal arch (TPA) with the use of a new typodont simulation system, the Calorific machine system, which was designed to observe the whole process of tooth movement. The maxillary right first molar was used for the anchor tooth and the maxillary left first molar was used for the mesial-in rotated tooth, and the angle of rotation was increased to 20, 40, and 60. A passive precision TPA was fabricated and then activated by bending the left arm to 20, 40, and 60. Each experiment was repeated five times under the same conditions and analyzed by ANOVA and Tucky's Studentized Range (HSD) test. In the occlusal plane, when the bending angle of precision TPA was increased, the mesiobuccal cusp of the rotated molar moved more buccally(p<0.001) and less distally (p<0.001) while the distolingual cusp moved in the mesiopalatal direction. In the sagittal plane, the palatal roots of the derotated molar moved mesially (p<0.001). In the traverse plane, the derotated molar showed slight extrusion (p<0.001). The upper right first molar, which was used as an anchor tooth, showed clinically insignificant movement across all three planes.

Propofol treatment modulates neurite extension regulated by immunologically challenged rat primary astrocytes: a possible role of PAI-1

고현명,주소현,이성훈,김희진,이승현,정재훈,류종훈,김정민,구본녀,신찬영 대한약학회 2015 Archives of Pharmacal Research Vol.38 No.4

Propofol, a widely used anesthetic, regulatesneurological processes including neurotoxicity, neuroprotection,glial activation, synaptic plasticity and neuronalmaturation. Tissue plasminogen activator/tissue plasminogenactivator inhibitor-1 (tPA/PAI-1) in CNS acts as aneuromodulator regulating synaptic plasticity, neuriteoutgrowth, seizure spreading and cell survival. Here, weinvestigated the effects of propofol on tPA/PAI-1 systemusing cultured neurons and astrocytes and their role in theregulation of neurite extension. Cultured rat primaryastrocytes were treated with propofol (1–10 lM) and LPS(10 ng/ml). The expression of functional tPA/PAI-1 wasexamined by casein zymography, Western blot and RTPCR. Alternatively, culture supernatants were added tocultured rat primary neuron to investigate the effects onneurite extension. Propofol alone did not affect tPA activityin rat primary cortical neuron. Similarly, propofol alonechanged neither tPA nor PAI-1 activity in rat primaryastrocytes. In immunologically challenged situation usingLPS, propofol synergistically increased expression of PAI-1 in rat primary astrocytes without affecting tPA expressionin a manner dependent on MAPKs activation. Increased expression of PAI-1 reduced tPA activity in LPSplus propofol-treated rat primary astrocytes. Consistentwith the critical role of tPA activity in the regulation ofneurite extension (Cho et al. 2013), the diminished tPAactivity in astrocyte culture supernatants resulted indecreased neurite extension when administered to culturedrat primary cortical neuron. The results from the presentstudy suggest that propofol, especially in immunologicallychallengedsituation, dysregulates tPA/PAI-1 system inbrain. Whether the dysregulated tPA/PAI-1 activityadversely affects neural differentiation as well as regenerationof neuron in vivo should be empirically determined inthe future.

Biocatalyst including porous enzyme cluster composite immobilized by two-step crosslinking and its utilization as enzymatic biofuel cell

Chung, Yongjin,Christwardana, Marcelinus,Tannia, Daniel Chris,Kim, Ki Jae,Kwon, Yongchai Elsevier Sequoia 2017 Journal of Power Sources Vol. No.

<P><B>Abstract</B></P> <P>An enzyme cluster composite (TPA/GOx) formed from glucose oxidase (GOx) and terephthalaldehyde (TPA) that is coated onto polyethyleneimine (PEI) and carbon nanotubes (CNTs) is suggested as a new catalyst ([(TPA/GOx)/PEI]/CNT). In this catalyst, TPA promotes inter-GOx links by crosslinking to form a large and porous structure, and the TPA/GOx composite is again crosslinked with PEI/CNT to increase the amount of immobilized GOx. Such a two-step crosslinking (i) increases electron transfer because of electron delocalization by π conjugation and (ii) reduces GOx denaturation because of the formation of strong chemical bonds while its porosity facilitates mass transfer. With these features, an enzymatic biofuel cell (EBC) employing the new catalyst is fabricated and induces an excellent maximum power density (1.62 ± 0.08 mW cm<SUP>−2</SUP>), while the catalytic activity of the [(TPA/GOx)/PEI]/CNT catalyst is outstanding. This is clear evidence that the two-step crosslinking and porous structure caused by adoption of the TPA/GOx composite affect the performance enhancement of EBC.</P> <P><B>Highlights</B></P> <P> <UL> <LI> Porous [TPA/GOx]/PEI/CNT catalyst is suggested by using two-step crosslinking. </LI> <LI> The catalyst increases electron transfer by electron delocalization by π conjugation. </LI> <LI> The catalyst curtails enzyme denaturation by strong chemical bonding. </LI> <LI> The catalyst induces superior EBC performance. </LI> <LI> Terephthalaldehyde (TPA) promotes inter-GOx links to form large and porous structure. </LI> </UL> </P> <P><B>Graphical abstract</B></P> <P>[DISPLAY OMISSION]</P>

REDUCED LEVEL OF ATF IS CORRELATED WITH TRANSCRIPTIONAL REPRESSION OF DNA TOPOISOMERASE Ⅱα GENE DURING TPA-INDUCED DIFFERENTIATION OF HL-60 CELLS

Lim, Kyu,Lee, Jong-In,Yun, Kyung-Ah,Son, Mee-Young,Park, Jong-Il,Yoon, Wan-Hee,Hwang, Byung-Doo 충남대학교 생물공학연구소 1999 생물공학연구지 Vol.7 No.-

DNA topoisomeraseⅡ is a marker for the proliferation state of mammalian cells in culture, and the protein levels are markedly higher in exponentially growing cells than quiescent cells and can be downregulated by growth of the cells at high density and serum starvation. Correlation between ATF and TPA-repressed DNA topoisomeraseⅡα (Topo Ⅱα) mRNA has been investigated during TPA-induced differentiation of HL-60 cells. Topo Ⅱα mRNA and unknotting activity were reduced at 24 hours in TPA-treated HL-60 cells. The level of Topo Ⅱα mRNA and the activity were gradually decreased in proportion to the concentration of TPA. Two DNA-protein complexes were formed by DNA mobility shift assay when ATF-binding site was incubated with nuclear extract prepared from TPA-free HL-60 cells, and the amount of ATF was vanished after TPA treatment. TPA-repressed TopoⅡα mRNA and ATF levels were partially restored after pretreatment of staurosporin. These results suggest that the reduced level of ATF may be important to the transcriptional repression of TopoⅡα gene during TPA-induced differentiation in HL-60 cells and related to protein kinase C signal pathway.