缝隙连接蛋白43与神经胶质瘤

缝隙连接蛋白43(connexin43,Cx43)是中枢神经系统最丰富的缝隙连接蛋白(connexins,Cxs),主要在星形胶质细胞中表达;而在星形胶质细胞瘤(中枢神经系统最常见的肿瘤)中,Cx43表达量减少或缺失。许多研究表明,Cx43表达减少会促进神经胶质瘤细胞增殖。转染Cx43 cDNA于胶质瘤细胞后,其恶性表型逆转、生长速度减慢、致瘤性明显降低;胶质瘤细胞迁移能力却增强。除了Cx43形成的缝隙连接(gap junction,GJ)对神经胶质瘤有影响外,Cx43形成的半通道(hemichannels,HCs)及Cx43 C末端也起着重要的作用。     

防治血管再狭窄的新靶点——缝隙连接蛋白43

缝隙连接是介导相邻细胞间离子和小分子信号物质直接交换的跨膜通道,血管平滑肌细胞中主要表达连接蛋白43(connexin43,CX43)。研究表明,CX43表达上调与平滑肌细胞的增殖、迁移及细胞外基质生成密切相关,促使血管损伤后新内膜生成,引起血管再狭窄,提示CX43可能成为血管性疾病治疗的新靶点。本文综述CX43与血管再狭窄之间关系及以CX43为靶点的药物和基因治疗的研究进展。     

连接蛋白Connexin43在鼻息肉中的表达

目的：研究连接蛋白43（Connextio 43,Cx43）在正常鼻黏膜和鼻息肉中的表达,探讨鼻息肉的发病机制。方法：采用免疫组织化学染色和平均光密度检测技术,对30例鼻息肉和10例正常鼻黏膜组中Cx43的表达进行分析。结果：连接蛋白Connexin 43主要表达于黏膜及黏膜下组织,正常鼻黏膜表达最明显（P〈0.01）,鼻息肉中表达相对于正常鼻黏膜中表达普遍减少。结论：连接蛋白Connexin 43在鼻黏膜和鼻息肉中均表达,且主要集中在黏膜和黏膜下组织,Cx43在鼻息肉中表达下调可能与鼻息肉的发生存在着某种密切的联系。     

The 2,2',4,4',5,5'-Hexachlorobiphenyl-Enhanced Degradation of Connexin 43 Involves Both Proteasomal and Lysosomal Activities

One of the toxic effects of non-dioxin–like polychlorinatedbiphenyls (NDL-PCBs) is the acute inhibition of gap junctionalintercellular communication (GJIC), an event possibly associatedwith tumor promotion. The model NDL-PCB-2,2',4,4',5,5'-hexachlorobiphenyl(PCB 153)—induces a sustained GJIC inhibition in rat liverepithelial WB-F344 cells. As this effect might be related toderegulation of connexin 43 (Cx43) synthesis, trafficking, ordegradation, we investigated the impact of PCB 153 on theseevents. Although PCB 153 had no effect on Cx43 mRNA levels,it induced a gradual loss of Cx43 protein and significantlydecreased the amount of gap junction plaques in plasma membrane.PCB 153 contributed to extracellular signal–regulatedkinases 1 and 2 (ERK1/2)–dependent accumulation of hyperphosphorylatedCx43-P3 form, thus indicating that ERK1/2 activation by PCB153 might contribute to its effects on Cx43 internalizationor degradation. Inhibition of either proteasomes or lysosomeswith their specific inhibitors largely restored total Cx43 proteinlevels, thus suggesting that both proteasomes and lysosomesmay participate in the PCB 153–enhanced Cx43 internalizationand degradation. However, neither the proteasomal nor the lysosomalinhibitors restored normal GJIC or number/size of gap junctionplaques. Finally, PCB 153 also interfered with restoration ofgap junction plaques following the inhibition of Cx43 transportto plasma membrane. Taken together, multiple modes of actionseem to contribute to downregulation of Cx43 in PCB 153–treatedrat liver epithelial cells. The enhanced degradation of Cx43,together with persistent inhibition of GJIC, might contributeto tumor-promoting effects of NDL-PCBs.     

Silver nanoparticles up-regulate Connexin43 expression and increase gap junctional intercellular communication in human lung adenocarcinoma cell line A549

Abstract

Silver nanoparticles (Ag NPs) are increasingly being used in wound dressings, medical settings, and various household products due to their unique properties and antimicrobial activity. Despite the widespread use of Ag NP products, the molecular mechanisms underlying the biological effects of Ag NPs remain unclear. Gap junctional intercellular communication (GJIC), formed by the connexin protein family, plays a critical role in the maintenance of tissue and organ homeostasis. This study was undertaken to investigate the effects of well characterized, PVP-coated Ag NPs (69 ± 3 nm) and silver nitrate on GJIC and connexin43 (Cx43) expression in human lung adenocarcinoma cell line A549. Our results showed that Ag NPs increased GJIC in A549 cells as assayed by dye transfer method. Western blot analysis showed that incubation of cells with Ag NPs significantly increased the expression of Cx43 protein. In addition, Ag NPs up-regulated expression of Cx43 mRNA in a dose-dependent manner. Silver nitrate failed to increase GJIC and the expression of Cx43 protein. It, however, increased Cx43 mRNA expression in A549 cells. Taken together, our results provide the first evidence that Ag NPs induced the increase of GJIC activity in A549 cells through up-regulation of Cx43 protein, suggesting that Cx43 and GJIC may be one of the targets for Ag NPs biological effects.     

丁苯酞对于星型胶质细胞炎症反应的保护作用及缝隙连接蛋白Cx43表达的影响

目的 研究丁苯酞（dl-3-n-butylphthalidle,NBP）对于脂多糖（lipopolysaccharide,LPS）诱导的星型胶质细胞（astrocytes,Ast）炎症反应的抑制作用和机制,同时观察其对于缝隙连接蛋白Cx43表达的影响。方法 Ast细胞采用梯度浓度的NBP处理,1 μg·mL^-1的LPS诱导炎症反应。试剂盒检测乳酸脱氢酶（lactate dehydrogenase,LDH）漏出量,CCK-8法检测细胞存活率,Griess法检测NO的释放水平,ELISA法检测培养基中炎症因子IL-1β、IL-6及TNF-α的表达水平。Western blotting法检测p65,p-IκB,Cx43的表达。免疫荧光染色法检测Cx43的分布。siRNA沉默Ast中Cx43的表达,LPS诱导炎症发生后,检测Ast的炎症因子释放以及细胞存活水平。结果 NBP可以降低Ast细胞中LDH的漏出,提高细胞存活率,抑制炎症因子的释放,同时抑制细胞中p65、p-IκB的表达。NBP可以同时抑制Ast中Cx43的表达。siRNA沉默Cx43后,Ast的炎症反应得到抑制。结论 丁苯酞可以抑制LPS诱导的Ast炎症反应,而Cx43与炎症反应的发生有关。丁苯酞抑制Ast炎症反应与抑制Cx43的表达有关。     

缝隙连接蛋白-43在糖尿病膀胱豚鼠模型中的表达及意义

目的 探讨缝隙连接蛋白-43（Cx43）在豚鼠糖尿病膀胱（DCP）中的表达情况及其意义。方法 50只豚鼠 随机分为DCP组（n=30）、枸橼酸钠组（n=10）、空白对照组（n=10）,DCP组豚鼠单次腹腔注射链脲佐菌素建立糖尿病 模型,利用尿动力学检测筛选出糖尿病膀胱豚鼠;枸橼酸钠组豚鼠单次腹腔注射枸橼酸钠溶液;空白对照组豚鼠单 次腹腔注射生理盐水;通过免疫组织化学染色法观察Cx43在3组膀胱逼尿肌中的定位及分布,采用Western blot技术 检测3组膀胱逼尿肌组织内Cx43蛋白的表达。结果 免疫组织化学染色结果显示,DCP组（筛选出DCP豚鼠共10只 纳入研究）Cx43特异性染色强度较其他2组明显降低,枸橼酸钠组和空白对照组阳性染色强度对比无明显差异。 Western blot检测发现,DCP组膀胱组织Cx43蛋白表达（0.52±0.02）低于空白对照组（0.68±0.02）和枸橼酸钠组（0.70± 0.03）（P＜0.01）,空白对照组与枸橼酸钠组膀胱组织Cx43蛋白表达差异无统计学意义（P＞0.05）。结论 逼尿肌组 织Cx43的表达降低在糖尿病膀胱的形成过程中起重要作用,可能成为研究糖尿病膀胱发病机制的新出发点。     

白藜芦醇能够修复离体缺血再灌注损伤大鼠心脏M3受体与间隙连接蛋白43间结构及功能性整合

为了探讨白藜芦醇是否能通过影响M₃受体和间隙连接蛋白43(Cx43)间结构及功能性整合发挥其抗心肌缺血再灌注损伤作用，应用免疫共沉淀、免疫印迹及免疫荧光技术研究白藜芦醇对M₃受体与Cx43间结构及功能性整合的影响。结合大鼠离体II导联心电图及心肌超氧化物歧化酶(SOD)、丙二醛(MDA)的检测观察白藜芦醇是否能恢复心肌缺血再灌注损伤。白藜芦醇能修复心肌缺血再灌注损伤所致的M₃受体与Cx43间结构及功能性整合的破坏及纠正Cx43表达异常。同时QRS波时限﹑SOD及MDA的改变也得到相应恢复。白藜芦醇能修复M₃受体与Cx43间结构及功能性整合而发挥抗缺血再灌注损伤作用。     

Connexin 43在神经原性逼尿肌过度活动中表达的实验研究

目的 研究Connexin 43在神经原性逼尿肌中的表达情况.方法 选择神经原性逼尿肌过度活动(neurogenic detrusor overactivity,NDO)患者16例为Ⅰ组,压力性尿失禁(stress urinary incontinence,SUI)膀胱逼尿肌功能正常患者10例为Ⅱ组,取两组逼尿肌标本,利用免疫组织化学和RT-PCR技术检测两组逼尿肌细胞中Connexin43及其蛋白表达变化情况.结果 免疫组织化学及RT-PCR技术检测结果显示,Ⅰ组逼尿肌细胞中Connexin 43及其蛋白含量明显高于Ⅱ组(P＜0.05).结论 神经原性逼尿肌过度活动逼尿肌中Connexin 43基因表达增加,可能与神经原性逼尿肌过度活动的发生有密切关系.     

以siRNA表达载体稳定抑制缝隙连接蛋白43表达的睾丸间质细胞和睾丸支持细胞系的建立

目的构建Cx43-siRNA真核表达载体,获得连接蛋白43(connexin43,Cx43)被长期稳定抑制的睾丸间质细胞(TM3细胞)系和睾丸支持细胞(TM4细胞)系,为研究Cx43及其形成的细胞缝隙连接(gap junction,GJ)在睾丸组织中的作用提供有用模型。方法设计合成3对针对Cx43的短发夹样siRNA的DNA模板序列,定向连接到siRNA真核表达载体pSilencerTM2.1-U6neo上,通过测序鉴定后以脂质体法瞬时转染睾丸支持细胞,以Western blot方法检测Cx43蛋白表达水平,筛选出最有效的干扰序列,再将之分别转染睾丸间质细胞和睾丸支持细胞,G418筛选出能稳定表达siRNA的细胞系,以"Parachute"荧光传递示踪法检测细胞缝隙连接功能。结果 Western blot结果显示,第3对干扰序列对Cx43表达抑制效果最佳,以表达该序列的质粒稳定转染的TM3细胞和TM4细胞上Cx43蛋白表达水平均明显降低;荧光传递示踪法检测表明,两种细胞系的GJ功能均被明显抑制。结论以Cx43-siRNA真核表达载体稳定转染的方法能长期干扰TM3和TM4细胞上Cx43的表达,并抑制由其形成的GJ功能。     

在Hs578T乳腺癌细胞由Cx43组成的细胞缝隙连接对阿霉素细胞毒性的影响

目的探讨乳腺癌细胞Hs578T中缝隙连接蛋白43(connexin43,Cx43)的表达,以及由其形成的缝隙连接(gapjunction,GJ)对阿霉素(adriamycin,ADM)细胞毒性的影响。方法采用Western blot检测Hs578T细胞中Cx43表达水平;细胞免疫荧光法观察Hs578T细胞膜Cx43蛋白的表达;细胞接种荧光示踪法测定Hs578T细胞荧光传递功能;MTT法检测缝隙连接功能对ADM细胞毒性的影响。结果Hs578T细胞中天然表达Cx43,10μmol·L-1维甲酸(ratinoicacid,RA)处理细胞24 h后,细胞中Cx43蛋白表达水平增高,25μmol·L-1 oleamide和10μmol·L-118-α-GA分别处理细胞24 h后,细胞中Cx43蛋白表达水平降低;Hs578T细胞膜表面有Cx43蛋白表达;10μmol·L-1RA预处理细胞24 h,细胞间荧光传递功能增强(P<0.01),25μmol·L-1oleamide和10μmol·L-118-α-GA预处理细胞1 h,细胞间荧光传递功能降低(P<0.01);在高密度接种细胞(生长融合,有GJ形成),10μmol·L-1RA增强细胞GJ功能,6μmol·L-1ADM对细胞增殖抑制率明显增加(P<0.01),25μmol·L-1 oleamide或10μmol·L-1 18-α-GA抑制细胞GJ功能,6μmol·L-1ADM对细胞增殖抑制率降低(P<0.01),而在低密度接种细胞(生长未融合,无GJ形成)细胞增殖抑制率没有改变(P>0.05)。结论 Hs578T细胞天然表达Cx43蛋白,并且增强细胞间由Cx43形成的GJ功能,ADM的细胞毒性也增加;而抑制细胞GJ功能时,ADM的细胞毒性也相应降低。     

缝隙连接及其连接蛋白对血管加压素诱导的失血性休克大鼠血管收缩的作用研究

摘要：目的 观察缝隙连接（GJ）及其组成亚单位连接蛋白（Cx）在血管加压素（AVP）诱导失血性休克大鼠血管收缩中的作用。方法 采用失血性休克大鼠模型和缺氧培养血管平滑肌细胞（VSMC）,观察GJ阻断剂CBX和octanol、以及各Cx亚型反义寡核苷酸（AODN）对AVP诱导的血管收缩反应的影响,随后进一步观察参与AVP作用的Cx37和Cx43对AVP调节休克血管钙敏感性和缺氧VSMC内钙离子浓度的影响。结果 GJ阻断剂CBX和octanol明显抑制了AVP诱导的休克血管的收缩反应。在所有血管中表达的连接蛋白中,Cx37AODN和Cx43AODN明显抑制了AVP的血管收缩作用。进一步结果显示,Cx43AODN、而不是Cx37AODN,可拮抗AVP升高休克血管钙敏感性的作用。此外, AVP处理和干扰Cx37及Cx43对缺氧VSMC内钙离子浓度无明显影响。结论 缝隙连接在休克后AVP介导的血管收缩调节中有重要作用,Cx37和Cx43参与了这一过程,其中Cx43可能通过影响AVP介导的血管钙敏感性调节途径来发挥作用,而Cx37可能通过其它机制来参与AVP的血管调节作用。     

缝隙连接蛋白43在脑心综合症中的作用研究

目的检测缝隙连接蛋白43(connexin43,Cx43)在脑心综合症模型大鼠心室肌中的表达及分布,以探讨其与脑缺血后心律失常的关系。方法用线栓法栓塞大鼠右侧大脑中动脉制备脑心综合症模型,检测心电图,电镜下观察心肌细胞的亚显微结构,采用Westernblot和免疫组织化学的方法,检测心室肌中Cx43表达和分布的改变。结果电镜显示脑缺血后,心肌组织闰盘内桥粒和缝隙连接结构及分布改变,与正常对照组相比,Western blot和免疫组织化学结果均显示脑缺血后2h心室肌中Cx43的表达水平明显降低,而缺血后24h心室肌中Cx43的表达增加(P<0·05)。心电图显示,发生心律失常之前10个心动周期的心电QT间期较脑未梗死前延长(P<0·01)。结论大鼠右侧局灶性脑缺血所引起的心律失常可能与心室肌Cx43蛋白表达异常并导致的QT间期延长有关。     

白藜芦醇通过抑制膀胱癌细胞的糖代谢增强顺铂的抗肿瘤活性

目的 研究白藜芦醇增强顺铂对膀胱癌细胞的杀伤活性及机制。方法 MTT法和流式细胞术分别检测顺铂单独治疗及联合白藜芦醇治疗对膀胱癌细胞系T24的杀伤活性和凋亡诱导效应。将T24细胞用白藜芦醇和顺铂处理后,检测T24细胞对葡萄糖的摄取能力和乳酸及ATP的生成能力。将T24细胞用白藜芦醇和顺铂处理后,用Western blot方法检测T24细胞PKM2的表达水平。构建PKM2真核表达载体,检测PKM2表达载体对白藜芦醇联合顺铂杀伤T24细胞疗效的影响。结果 白藜芦醇单独治疗在体外对T24细胞的杀伤活性较低,但能显著增强顺铂对T24细胞的杀伤活性。白藜芦醇能显著减弱T24细胞对葡萄糖的摄取和乳酸及ATP的生成并下调T24细胞的PKM2表达水平。转染PKM2表达载体后,白藜芦醇对顺铂的协同抗肿瘤作用受到抑制。结论 白藜芦醇通过下调PKM2的表达抑制肿瘤细胞的糖代谢增强顺铂对膀胱癌细胞的杀伤活性。     

槲皮素对自发性高血压大鼠大脑中动脉张力与缝隙连接蛋白43表达的影响

目的研究槲皮素对自发性高血压大鼠(SHR)大脑中动脉张力的影响及其与缝隙连接蛋白43(Cx43)的关系。方法取WKY大鼠作为正常对照组(每日给予正常饮水摄食);按照体重将SHR大鼠随机分为2组:模型组(每日给予正常饮水摄食)和实验组(每日给予槲皮素50 mg·kg^-1,ig),每组10只。8周后,以血管张力测定法检测大鼠大脑中动脉对血管舒缩剂的反应性,用逆转录-聚合酶链反应和免疫印迹法检测大鼠大脑中动脉的Cx43 mRNA和Cx43蛋白表达水平。结果 U46619(血栓素A2类似物)收缩后,正常对照组、模型组和实验组的最大收缩百分比分别为(43. 15±3. 42)%,(100. 00±0. 00)%和(67. 38±4. 26)%; KCl收缩后,这3组大鼠的最大收缩百分比分别为(37. 23±5. 01)%,(100. 00±0. 00)%和(60. 23±4. 52)%,正常对照组和模型组比较或者模型组和实验组比较,这2种收缩的差异均有统计学意义(P <0. 05,P <0. 01)。硝普钠舒张后,这3组大鼠的最大舒张百分比分别为(85. 16±4. 23)%,(50. 25±3. 17)%和(75. 18±3. 06)%;吡那地尔舒张后,这3组大鼠的最大舒张百分比分别为(83. 25±4. 37)%,(53. 06±3. 53)%和(70. 23±4. 21)%,正常对照组和模型组比较或者模型组和实验组比较,这2种舒张的差异均有统计学意义(P <0. 05,P <0. 01)。这3组大鼠大脑中动脉的Cx43 mRNA表达量分别为0. 63±0. 12,1. 30±0. 17和0. 80±0. 15,正常对照组和模型组比较或者模型组和实验组比较,差异均有统计学意义(均P <0. 01);这3组大鼠大脑中动脉的Cx43蛋白表达趋势与mRNA结果一致。结论槲皮素可以通过抑制大脑中动脉的Cx43表达改善SHR大脑中动脉的张力异常。     

RETRACTED ARTICLE: Chemopreventive Potential of Resveratrol in Mouse Skin Tumors Through Regulation of Mitochondrial and PI3K/AKT Signaling Pathways

Purpose  To investigate the chemopreventive potential of resveratrol, a phytoalexin found in seeds and skin of grapes, berries and peanuts in 7,12 dimethyl benz(a)anthracene (DMBA) induced mouse skin tumorigenesis. Methods  Topical treatment of resveratrol was given to the animals 1 h prior to DMBA for 28 weeks. At the end of the study period, the skin tumors were dissected out and western blotting was carried out to examine the regulation of proteins involved in anti-tumorigenesis in response to resveratrol. Results  Chemopreventive properties of resveratrol were reflected by delay in onset of tumorigenesis, reduced cumulative number of tumors, and reduction in tumor volume. Results of the western blotting showed that resveratrol treatment increased the DMBA suppressed p53 and Bax while decreased the expression of Bcl-2 and Survivin. Further, resveratrol supplementation resulted in release of cytochrome C, caspases activation, increase in apoptotic protease-activating factor-1 (Apaf-1) as mechanism of apoptosis induction. Resveratrol was also found to inhibit skin tumorigenesis through regulation of Phosphatidylinositol-3-kinase (PI3K)/ and AKT proteins which are implicated in cancer progression because it stimulates proliferation and suppresses apoptosis. Conclusions  Based on the results we can conclude that resveratrol regulates apoptosis and cell survival in mouse skin tumors as mechanism of chemoprevention hence deserve to be a chemopreventive agent.     

Effect of hypaconitine combined with liquiritin on the expression of calmodulin and connexin43 in rat cardiac muscle in vivo

Objectives To study the effects of hypaconitine used alone and combined with liquiritin on calmodulin (CaM) expression and connexin43 (Cx43) phosphorylation on serine368 (Ser368), as well as to investigate the intervention of liquiritin on these hypaconitine‐induced effects. Methods Adult Wistar rats were orally administered hypaconitine (0.23, 0.69, 2.07 mg/kg per day), liquiritin (20 mg/kg per day), or hypaconitine (2.07 mg/kg per day) plus liquiritin (20 mg/kg per day) for seven consecutive days. The mRNA expression levels of CaM and Cx43 in rat myocardial tissue were determined by real‐time quantitative PCR. The protein contents of CaM and phosphorylated Cx43 (Ser368) were determined by Western blot. Key findings The results indicated that the mRNA and protein expression levels of CaM were significantly decreased by hypaconitine used alone and combined with liquiritin. Although CaM mRNA expression level was inhibited by liquiritin, its protein expression level was upregulated. Meanwhile, although no obvious effect on Cx43 mRNA expression was observed after the drug administration, the phosphorylation level of Cx43 (Ser368) was significantly inhibited. Furthermore, the coadministration of hypaconitine and liquiritin significantly reduced hypaconitine‐induced inhibitory action on Cx43 (Ser368) phosphorylation. Conclusions The study indicated that hypaconitine could inhibit CaM expression and Cx43 (Ser368) phosphorylation, and liquiritin could interfere with this kind of effect by synergistically inhibiting CaM expression and by antagonizing Cx43 (Ser368) dephosphorylation induced by hypaconitine.     

Connexin 43 mediates changes in protein phosphorylation in HK-2 cells during chronic cadmium exposure

Connexin 43 (Cx43) is believed to play a role in the mechanisms of toxicity of many chemical species, include cadmium (Cd). In this study, human renal proximal tubule (HK-2) cells were exposed to Cd (1 μM, 10 days). Of the 584 protein residues detected using a Phospho Explorer antibody microarray (PEX100), more than half changed their levels of phosphorylation after chronic Cd exposure. Cx43 siRNA attenuated Cd-induced apoptosis and inhibited proliferation, while also attenuating changes in the levels of phosphorylation of many protein residues. According to DAVID Bioinformatics Resources analysis and KEGG PATHWAY database, AKT signal pathway may be the important one. Focusing on the AKT pathway confirmed that Cx43 mediated increased levels of p-PTEN^{Ser380/Ser382/Thr383} and decreased levels of p-AKT^Thr308, p-AKT^Tyr326, p-ASK1^Ser83, and p-p27^Thr187, thereby possibly contributing to the Cd-induced apoptosis and inhibited proliferation. These results suggested that AKT pathway was the dominant pathway involved in Cx43-mediated chronic Cd toxicity.     

Nafenopin causes protein kinase C-mediated serine phosphorylation and loss of function of connexin 32 protein in rat hepatocytes without aberrant expression or localization.

The characteristics and mechanism of the inhibition of connexin-mediated gap junctional communication by the non-genotoxic rodent hepatocarcinogen, nafenopin, has been studied in rat hepatocytes. Nafenopin caused a time- and concentration-dependent inhibition of dye coupling in hepatocytes as assessed by transfer of microinjected lucifer yellow. A half-maximum inhibitory effect of nafenopin occurred at approximately 50 microM, which was not cytotoxic. The inhibitory effect was reversible since a significant recovery of communication was observed 3 h after removal of the chemical. The protein kinase inhibitor G?6976 prevented the inhibition of dye coupling, but a tyrosine kinase inhibitor (genistein) did not. Connexin 32 and 26 protein expression, as assessed by immunoblotting, was similar in nafenopin-treated hepatocytes compared to controls, with the exception that in a 10-h culture with nafenopin, the level of connexin 26 was elevated compared to controls. Immunohistochemistry indicated that the localization of plaques containing connexin 32 was not affected in hepatocytes by nafenopin. Immunoprecipitated connexin 32 was, however, detected by an anti-phosphoserine antibody following nafenopin treatment, but not in controls. This serine phosphorylation was prevented in the presence of G?6976. The results give further support for a role of protein kinase C in the post-translational inactivation of connexin 32 function in rat hepatocytes by nafenopin.