过敏性哮喘患者高迁移率族蛋白1、Th1/Th2细胞亚群及相关细胞因子表达分析

目的：分析高迁移率族蛋白-1（high mobility group protein-1，HMGB-1）、辅助T淋巴细胞（Th）亚群以及相关细胞因子在不同严重程度的过敏性支气管哮喘患者中的表达意义。方法：选择98例轻中重度过敏性哮喘患者和30例同期体检的健康者（对照组），采用流式细胞术分析患者以及健康者外周血CD4⁺干扰素-γ⁺（IFN-γ⁺）T细胞、CD4⁺白细胞介素-4⁺（IL-4⁺）T细胞百分比。通过酶联免疫吸附测定法（ELISA）检测血清中HMGB-1、Th1细胞因子（IFN-γ）和Th2细胞因子（IL-4、IL-5）的表达水平。采用Spearman分析HMGB-1、Th1/Th2细胞亚群及其相关细胞因子与肺功能的相关性。结果：所有哮喘患者HMGB-1、CD4⁺IL-4⁺T细胞百分比、IL-4、IL-5均高于对照组；重度组患者HMGB-1、CD4⁺IL-4⁺T细胞百分比、IL-4、IL-5高于轻、中度患者。所有患者CD4⁺IFN-γ⁺T细胞百分比、IFN-γ表达均低于对照组；重度患者CD4⁺IFN-γ⁺T细胞百分比和IFN-γ表达低于轻、中度患者。HMGB-1、IL-4水平与肺功能负相关，IFN-γ水平与肺功能正相关。结论：HMGB-1和IFN-γ、IL-4可作为评价支气管哮喘严重程度的有效指标。     

低强度超声对离体大鼠膀胱平滑肌收缩的影响

目的 观察低强度脉冲超声(LIPUS)对离体大鼠膀胱平滑肌收缩的影响。方法 将32只SD雌性大鼠的32段离体膀胱肌条随机分为超声辐照组、超声辐照+尼莫地平组、超声辐照+低分子肝素组及对照组,每组各8段。采用BL-410F生物机能实验系统测量并记录超声辐照组、超声辐照+尼莫地平组、超声辐照+低分子肝素组在LIPUS辐照前、后的收缩曲线,并比较3组膀胱平滑肌条收缩频率、幅度。对超声辐照组LIPUS辐照后和对照组膀胱平滑肌进行组织病理学观察。结果 与LIPUS辐照前比较,辐照后超声辐照组的收缩频率和幅度均明显增加(P均<0.05),超声辐照+尼莫地平组的收缩频率和幅度均明显降低(P均<0.05),超声辐照+低分子肝素组的收缩频率和幅度均无明显变化(P均>0.05)。组织病理学显示超声辐照组膀胱平滑肌细胞与对照组比较无明显差异。结论 LIPUS辐照可激活膀胱平滑肌细胞膜L-型钙通道,继而引起膀胱平滑肌的收缩增强。     

外周血T淋巴细胞亚群水平对结直肠癌患者术后复发的影响及预测价值分析

目的 探讨外周血T淋巴细胞亚群水平对腹腔镜根治术后结直肠癌（CRC）患者术后复发的影响,并分析其预测价值。方法 纳入2016年1月－2017年1月该院行腹腔镜根治术治疗的CRC患者120例,分为复发组（n = 96）和未复发组（n = 24）,检测患者术后第１天外周血中CD3⁺、CD4⁺、CD8⁺和CD4⁺/CD8⁺的含量,采用两独立样本t检验分析两组间差异,Logistic回归模型分析各指标与复发的关联,受试者工作特征曲线（ROC）分析其在预测CRC术后复发中的价值。结果 术后复发组CD3⁺、CD4⁺和CD4⁺/CD8⁺水平明显低于未复发组（P < 0.05）,CD8⁺水平高于未复发组（P < 0.05）。Logistic回归分析表明,血清中CD8⁺细胞水平（O = 1.14 > 1,95%CI：1.05～1.23）是影响结CRC术后复发的独立危险因素,而CD3⁺（O = 0.82,95%CI：0.75～0.90）、CD4⁺（O = 0.91,95%CI：0.85～0.98）和CD4⁺/CD8⁺（O = 0.20,95%CI：0.06～0.59）是影响CRC术后复发的保护性因素。ROC曲线表明,CD3⁺、CD4⁺、CD8⁺和CD4⁺/CD8⁺水平对术后复发具有一定的预测价值。结论 外周血中CD3⁺、CD4⁺、CD8⁺和CD4⁺/CD8⁺的水平与复发有关,是CRC患者术后复发的独立影响因素,与预后相关,对术后复发具有一定的预测价值。     

慢性乙型肝炎患者CD4+Th细胞中LSD1表达及其作用

目的： 探讨慢性乙型肝炎（乙肝）患者外周血CD4⁺Th细胞内赖氨酸特异性组蛋白去甲基化酶1（histone lysine specific demethylase 1，LSD1）表达水平与Th1/Th2平衡的关系，初步探讨LSD1介导的组蛋白修饰对慢性乙肝患者CD4⁺Th细胞分化的作用。方法： 选择2017年6月至12月在南通大学附属医院感染科住院的慢性乙肝患者65例，分为丙氨酸转氨酶（ALT）高水平组与低水平（≤4×ULN）组，HBV-DNA高载量（≥10⁶拷贝/mL）组与低载量（<10⁶拷贝/mL）组。以同期在该院体检的30例健康人为对照。采用密度梯度离心法分离外周血单个核细胞，免疫磁珠法分选CD4⁺Th细胞；提取CD4⁺Th细胞总蛋白，以Western印迹法检测LSD1表达水平。采用ELISA法检测血清中干扰素γ（IFN-γ）、白细胞介素4（IL-4）含量；用微粒子化学发光法检测血清乙型肝炎表面抗原（HBsAg）含量；用实时定量PCR法检测血清HBV-DNA载量。结果： 慢性乙肝组患者外周血CD4⁺Th细胞中LSD1表达水平高于健康人（0.52±0.21 vs 0.28±0.09，t=-7.49，P<0.001）。慢性乙肝患者中，CD4⁺Th细胞中LSD1表达量在ALT高水平组（n=38）低于低水平组（n=27，0.39±0.18 vs 0.64±0.16；t=-5.79，P<0.001），在HBV-DNA高载量组（n=32）高于HBV-DNA低载量组（n=33，0.69±0.08 vs 0.35±0.16；t=10.80，P<0.001）。与健康人相比，慢性乙肝组血清IFN-γ含量降低、IL-4含量增高、IFN-γ/IL-4比值减小（P<0.05）。慢性乙肝患者外周血CD4⁺Th细胞中LSD1表达量与其血清ALT水平（r=-0.590）、IFN-γ水平（r=-0.379）及IFN-γ/IL-4（-0.285）负相关（P<0.01），与HBV-DNA载量正相关（r=0.880，P<0.001），与血清IL-4水平无相关性（r=0.169，P=0.102）。结论： LSD1高表达可能是慢性乙肝患者体内Th1/Th2失衡、Th1反应水平下降的原因之一，并导致机体清除HBV能力减弱或抑制HBV复制能力减弱。     

超声引导下两种神经阻滞方式对胸腔镜下肺叶切除术患者炎症反应及免疫功能的影响

目的 比较超声引导下两种神经阻滞方式对胸腔镜下肺叶切除术患者炎症及免疫功能的影响,以期为该类患者的镇痛治疗提供参考。方法 选择2020年1月－2021年7月该院114例行胸腔镜下肺叶切除术的患者,排除1例脱落病例,最终纳入113例患者,根据麻醉方式的不同,将其分为A组（n = 57）和B组（n = 56）。两组患者均在超声引导下进行阻滞,A组采用胸椎旁神经阻滞,B组采用竖脊肌平面阻滞。比较两组患者围手术期情况、炎症反应、应激反应和免疫反应相关指标,以及不良反应发生率。结果 两组患者围手术期相关指标比较,差异均无统计学意义（P > 0.05）;两组患者术毕（T₁）和术后24 h（T₂）白细胞介素-10（IL-10）、白细胞介素-6（IL-6）、白细胞介素-1β（IL-1β）、皮质醇（Cor）、去甲肾上腺素（NE）及CD8⁺水平均较术前（T₀）明显升高（P < 0.05）;两组患者CD3⁺、CD4⁺及CD4⁺/CD8⁺水平较T₀时点明显降低（P < 0.05）,且A组明显高于B组（P < 0.05）;A组不良反应发生率明显低于B组（P < 0.05）。结论 胸腔镜下肺叶切除术患者采用超声引导下胸椎旁神经阻滞,有助于减轻机体应激及炎症反应,对患者免疫功能的影响较轻,安全性较高。     

彩色多普勒超敏血流显像技术显示孕11~13+6周胎儿心脏轴

目的 探讨彩色多普勒超敏血流（HD flow）显像技术在11~13⁺⁶周胎儿心脏轴显像中的价值。方法 采用灰阶超声及彩色多普勒HD flow显像技术对197胎孕11~13⁺⁶周胎儿进行心脏轴显像,比较两种方法对不同孕周胎儿心脏轴的显示率;采用两种方法测量55胎孕13~13⁺⁶周胎儿的心脏轴值,并进行比较。结果 孕11~13⁺⁶周,灰阶超声显像和HD flow显像对心脏轴的显示率分别为67.01%（132/197）、85.28%（168/197）,差异有统计学意义（P<0.01）;孕11~11⁺⁶周,显示率分别为32.39%（23/71）、69.01%（49/71）,差异有统计学意义（P<0.01）;孕12~12⁺⁶周,显示率分别为78.26%（54/69）、92.75%（64/69）,差异有统计学意义（P<0.01）;孕13~13⁺⁶周,两种方法显示率均为96.49%（55/57）。灰阶超声显像和HD flow显像测量心脏轴值分别为（45.34±3.99）°、（43.62±3.33）°,差异有统计学意义（t=7.11,P<0.01）。灰阶超声测得数值离散度较大,HD flow显像测得数值相对集中。结论 与灰阶超声显像相比,彩色多普勒HD flow显像可提高孕11~12⁺⁶周胎儿心脏轴显示率,且可降低13~13⁺⁶周胎儿心脏轴值的测量误差。     

针刺合谷穴后前额叶脑功能血氧水平依赖信号与神经递质谷氨酸及谷氨酰胺复合物的关系

目的 观察针刺右侧合谷穴时前额叶血氧水平依赖（BOLD）信号、较正后谷氨酸谷氨酰胺复合物（GLx⁺）浓度及其相关性。方法 对76名健康受试者采集静息及刺激[手针及纤毛机械刺激针（纤毛针）]合谷穴时BOLD功能MRI（fMRI）及磁共振波谱（MRS）数据,分析BOLD信号定量值与Glx⁺浓度的相关性。结果 手针较纤毛针针刺激发更多负激活,以前额叶内侧皮层（mPFC）及前扣带回（ACC）为著。2组BOLD信号定量值差异和Glx⁺浓度差异均无统计学意义（P均>0.05）。BOLD定量信号与Glx⁺浓度均无明显相关（P均>0.05）,刺激前后Glx⁺浓度差异均无统计学意义（P均>0.05）。结论 手针针刺合谷穴诱发边缘叶-旁边缘叶-新皮层网络负激活效应,手针和纤毛针刺激前后Glx⁺浓度多无明显变化;BOLD信号定量值与Glx⁺浓度无明显相关。     

基于孕11~13+6周不同颈项透明层厚度增加标准建立模型筛查高风险人群21-三体综合征

目的 观察11~13⁺⁶孕周21-三体综合征胎儿超声表现，探讨应用不同颈项透明层（NT）增厚标准建立的模型针对高风险人群筛查11~13⁺⁶孕周21-三体综合征胎儿的效能。方法 回顾性分析经胎儿染色体检查确诊为21-三体综合征的106胎（阳性组）及染色体正常（阴性组）1 391胎孕11~13⁺⁶周胎儿。记录2组超声标记出现情况。采用不同NT增厚标准（NT>第95百分位数、NT≥3.0 mm及NT≥3.5 mm）建立筛查21-三体综合征胎儿回归方程，评价其一致性及筛查效能。结果 阳性组72胎（72/106，67.92%）出现超声标记，以NT增厚（>第95百分位数）[70/106（66.04%）]最多见；阴性组89.58%（1 246/1 391）胎儿超声未见异常。以NT>第95百分位数定义NT增厚，筛查胎儿21-三体综合征的敏感度为66.04%，高于以NT≥3.0 mm（χ²=18.05）及NT≥3.5 mm（χ²=23.04）为标准（P均<0.01）。Logistic回归分析显示，孕妇高龄、胎儿NT增厚、鼻骨缺失及胎儿水肿4项指标为筛查胎儿21-三体综合征的有效标记（P均<0.05）。以不同NT增厚标准制定筛查21-三体综合征回归方程，以NT>第95百分位数与NT≥3.0 mm为NT增厚标准制定方程的拟合优度好（χ²=4.45、0.83，P=0.11、0.36），其筛查21-三体综合征胎儿敏感度、特异度及AUC分别为66.00%、90.50%、0.78和50.00%、92.80%及0.70（P均<0.05）。结论 以NT>第95百分位数为胎儿NT增厚标准制定的回归方程筛查高风险人群11~13⁺⁶周胎儿21-三体综合征的效能较佳。     

腹腔镜下胃癌D2根治术联合射频消融应用于胃癌肝转移的临床疗效

目的 探讨腹腔镜下胃癌D2根治术联合射频消融肝转移瘤应用于胃癌肝转移的临床疗效。方法 选取该院2017年1月－2020年1月收治的可切除胃癌肝转移需行胃癌D2根治术联合射频消融的患者62例,随机分为对照组和观察组,每组各31例。对照组采用开腹胃癌D2根治术联合射频消融治疗,观察组采用腹腔镜下胃癌D2根治术联合射频消融治疗。比较两组患者手术指标、血清肿瘤标志物指标、免疫相关指标和并发症情况。结果 对照组术中出血量多于观察组,肠功能恢复时间、住院时间明显长于观察组（P < 0.05）,手术时间短于观察组（P < 0.05）;术后两组患者血清甲胎蛋白（AFP）、癌胚抗原（CEA）水平降低（P < 0.05）,观察组血清AFP、CEA水平均低于对照组（P < 0.05）;术后两组患者CD4⁺、CD8⁺、CD4⁺/CD8⁺水平均较术前降低（P < 0.05）,观察组CD4⁺、CD4⁺/CD8⁺水平明显高于对照组（P < 0.05）;观察组并发症发生率12.90%低于对照组41.94%（P < 0.05）。结论 腹腔镜下胃癌D2根治术联合射频消融治疗胃癌肝转移有利于患者术后康复,可明显改善血清肿瘤标志物水平及机体免疫功能,降低围术期并发症发生率。     

艾滋病合并消化道溃疡的内镜下表现

目的 探讨艾滋病（AIDS）合并不同类型消化道溃疡的内镜下特征。方法 收集2018年5月－2021年5月于该院消化科住院的20例AIDS合并消化道溃疡的患者的临床病例资料，总结归纳其内镜下表现和病理特征。结果 所有病例中发现：食管溃疡4例，胃溃疡3例，回盲部溃疡3例，回肠末端溃疡2例，结肠溃疡1例，直肠溃疡7例。病理提示：放线菌感染1例，结核杆菌感染2例，真菌感染1例，腺癌1例，鳞状细胞癌1例，非霍奇金淋巴瘤2例，慢性炎症12例。人类免疫缺陷病毒（HIV）核糖核酸（RNA）阳性者7例，HIV RNA值为1.40×10²～3.57×10⁵ IU/mL，平均（2.81±8.97）×10⁴ IU/mL。17例患者获得了CD4⁺的数据。其中，CD4⁺ < 250/μL者7例，CD4⁺ > 250/μL者10例。合并梅毒血清学阳性2例，血清巨细胞病毒（CMV）阳性1例。外周血HIV RNA阳性和CD4⁺ T细胞数量，与消化道恶性肿瘤（腺癌、鳞癌和非霍奇金淋巴瘤）（P = 0.268，P = 0.315）、特异性病原体感染（结核杆菌和放线菌）（P = 0.359，P = 0.621）以及普通炎性溃疡（P = 0.549，P = 0.058）无明显相关性。结论 HIV感染者的消化道溃疡病因与正常人群的溃疡病因明显不同，常由机会性感染或恶性肿瘤导致。如HIV感染患者因吞咽困难、胸骨后疼痛或腹痛至消化科门诊就诊，临床医师需警惕并发胃肠道疾病的可能，及时给予患者胃肠镜检查。     

Neuroprotective Effect of Low-Intensity Pulsed Ultrasound on the Mouse MPTP/MPP+ Model of Dopaminergic Neuron Injury

Ultrasound mediated neuromodulation has been demonstrated to a safe treatment strategy in the field of neuroscience. In this study, low-intensity pulsed ultrasound (LIPUS) was used to treat Parkinson's disease (PD) models induced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and 1-methyl-4-phenylpyridinium (MPP⁺) to explore the possibility of ultrasound neuroprotective effect on PD. The results demonstrated that LIPUS treatment can attenuate the central neurotoxicity of MPTP in mice, reduce the loss of tyrosine hydroxylase positive neurons in the substantia nigra pars compacta and decrease the apoptosis in the section of substantia nigra. The movement and balance dysfunctions in PD mice were improved with LIPUS treatment. In addition, we demonstrated that LIPUS can inhibit the decreased activity and increased apoptosis of dopaminergic neurons induced by MPP⁺, restrain the accumulation of reactive oxygen species (ROS) and decrease of mitochondrial membrane potential caused by MPP⁺. Moreover, LIPUS stimulation alone did not cause any cytotoxicity and tissue damage in our study. Taken together, the protective and regulatory effects of LIPUS on dopaminergic neurons make it possible as a new, safe and noninvasive treatment for PD.     

Neuroprotective Effect of Low-Intensity Pulsed Ultrasound Against MPP+-Induced Neurotoxicity in PC12 Cells: Involvement of K2P Channels and Stretch-Activated Ion Channels

Parkinson's disease is the second most common neurodegenerative disease. It is characterized by the loss of dopaminergic neurons in the substantia nigra pars compacta. 1-Methyl-4-phenylpyridinium (MPP⁺) is a dopaminergic neuronal toxin that is widely used in constructing Parkinson's disease models in vitro. Low-intensity pulsed ultrasound (LIPUS) is a non-invasive therapeutic approach that has neuromodulation and neuroprotective effects in the central neural system; however, whether LIPUS can provide protection for dopaminergic neurons against MPP⁺-induced neurocytotoxicity remains unknown. In this study, we found that pre-treatment with LIPUS (1 MHz, 50 mW/cm², 20% duty cycle and 100-Hz pulse repetition frequency, 10 min) inhibited MPP⁺-induced neurotoxicity and mitochondrial dysfunction in PC12 cells. LIPUS decreased MPP⁺-induced oxidative stress by modulating antioxidant proteins, including thioredoxin-1 and heme oxygenase-1, and prevented neurocytotoxicity via the phosphoinositide 3-kinase (PI3K)-Akt and ERK1/2 pathways. Furthermore, these beneficial effects were attributed to the activation of K2P channels and stretch-activated ion channels by LIPUS. These data indicate that LIPUS protects neuronal cells from MPP⁺-induced cell death through the K2P channel- and stretch-activated ion channel-mediated downstream pathways. The data also suggest that LIPUS could be a promising therapeutic method in halting or retarding the degeneration of dopaminergic neurons in Parkinson's disease in a non-invasive manner.     

Antitumor effect of N-succinyl-chitosan nanoparticles on K562 cells

Polymer nanoparticles have become attractive for its prominent property recent years. In this paper, antitumor effects of N-succinyl-chitosan nanoparticles (NSCNP), a nanoparticles derivated from chitosan, were evaluated in K562 cells, including cell viability comparison, cell morphology analysis, DNA fragmentation detection, cell surface potential and mitochondrial membrane potential (MMP) measurement, intracellular ROS and Ca²⁺ concentration evaluation. Our results revealed NSCNP could inhibit the proliferation of K562 with an IC₅₀ of 14.26 μg/ml (24 h); decrease the zeta potential; disrupt the mitochondrial membrane potential; increase ROS generation and Ca²⁺ concentration. Cytomorphology study and DNA fragment analysis reveal characteristics of apoptosis and necrosis, indicating that the antitumor effect of NSCNP achieved by necrosis and apoptosis induction in K562 cells.     

Retracted Article: Knockdown of NEAT1 ameliorated MPP+-induced neuronal damage by sponging miR-221 in SH-SY5Y cells

Long noncoding RNAs (lncRNAs) have recently attracted increasing attention for their involvement in a wide variety of human neurodegenerative diseases, including Parkinson''s disease (PD). The purpose of the present study was to investigate the functional role and underlying mechanism of NEAT1 in PD. qRT-PCR was used to assess the expression of NEAT1 and miR-221, and the expression levels of Bcl-2 and Bax were detected by western blot. Cell viability and apoptosis were determined by CCK-8 assay and flow cytometry, respectively. The changes of oxidative stress and neuroinflammation were evaluated by ELISA assay and qRT-PCR, respectively. The targeted interaction between NEAT1 and miR-221 was verified by dual-luciferase reporter assay and RNA immunoprecipitation assay. Our data supported that MPP⁺ treatment elevated NEAT1 expression in dose- and time-dependent manners in SH-SY5Y cells, and NEAT1 silencing relieved MPP⁺-induced suppression of cell viability and enhancement of cell apoptosis in SH-SY5Y cells. Moreover, NEAT1 silencing alleviated MPP⁺-induced promotion of oxidative stress and neuroinflammation in SH-SY5Y cells. NEAT1 directly targeted miR-221 and negatively regulated miR-221 expression. More importantly, miR-221 mediated the protective effect of NEAT1 knockdown, as evidenced by the restoration of cell viability, cell apoptosis, oxidative stress and neuroinflammation in MPP⁺-induced SH-SY5Y cells. In conclusion, our study suggested that NEAT1 silencing alleviated MPP⁺-induced neuronal damage by sponging miR-221 in SH-SY5Y cells, highlighting the role of NEAT1 as a potential molecular target for PD therapy.

Long noncoding RNAs (lncRNAs) have recently attracted increasing attention for their involvement in a wide variety of human neurodegenerative diseases, including Parkinson''s disease (PD).     

Allyl‐isatin suppresses cell viability,induces cell cycle arrest,and promotes cell apoptosis in hepatocellular carcinoma HepG2 cells

The anticancer effect of the newly synthesized isatin derivative, N‐allyl‐isatin (Allyl‐I), was evaluated in vitro with human hepatocellular carcinoma HepG2 cells. Cell viability was detected by cell counting kit‐8 (CCK8) assay. Acridine orange (AO)/ethidium bromide (EB) double staining was used to observe the cell morphology. Flow cytometry was used to assess the effects of Allyl‐I on the cell cycle, apoptosis rate, and mitochondrial membrane potential (MMP). Western blot analysis was performed to detect the influence of Ally1‐I on the expression of cytochrome c (cyt c), Bax, Bcl‐2, and cleaved caspase‐3. Allyl‐I significantly inhibited HepG2 cell viability in a time‐ and dose‐dependent manner. Allyl‐I can induce cell cycle arrest in HepG2 cells at the G2/M phase. Apoptotic nuclear morphological changes were observed after AO/EB double staining. Fluorescein isothiocyanate‐conjugated Annexin V (Annexin V‐FITC) and propidium iodide (PI) double staining showed that the apoptotic rates significantly increased in the presence of Allyl‐I. Rhodamine 123 staining indicated that Allyl‐I can decrease the MMP. Allyl‐I also altered the expression of mitochondrial apoptosis‐related proteins. Protein levels of cyt c and cleaved caspase‐3 were upregulated following Allyl‐I treatment. By contrast, the Bcl‐2/Bax ratio decreased. Results suggest that Allyl‐I suppresses cell viability, induces cell cycle arrest, and promotes cell apoptosis in HepG2 cells. Furthermore, the induction of apoptosis might be correlated with the mitochondrial pathway.     

Ultraviolet light-emitting diode irradiation induces reactive oxygen species production and mitochondrial membrane potential reduction in HL-60 cells

ObjectiveUltraviolet light-emitting diode (UV LED) irradiation at 280 nm has been confirmed to induce apoptosis in cultured HL-60 cells, but the underlying mechanisms remain unclear. This study aimed to investigate the effects of 280 nm UV LED irradiation on reactive oxygen species (ROS) production and mitochondrial membrane potential (MMP) in HL-60 cells.MethodsHL-60 cells were irradiated with 0, 8, 15, or 30 J/m² of 280 nm UV LED and incubated for 2 hours. The intracellular ROS levels were assessed using the fluorescent probe 2ʹ-7ʹ-dichlorodihydrofluorescein diacetate (DCFH-DA) and a fluorescence plate reader. MMP was determined by flow cytometry using 5,5ʹ,6,6ʹ-tetrachloro-1,1ʹ,3,3ʹ-tetraethylbenzimidazol-carbocyanine iodide (JC-1) staining. The apoptosis-related proteins Bax and Bcl-2 were evaluated by western blot.ResultsUV LED irradiation at 280 nm induced a dose-dependent increase in ROS production and loss of MMP, and it activated apoptosis at irradiation doses of 8 to 30 J/m². These results were consistent with a previous apoptosis study from the authors’ group.ConclusionEnhanced ROS production and mitochondrial depolarization are two distinct but interacting events, and both are involved in UV LED-induced apoptosis in HL-60 cells.     

盐酸右美托咪定调控HIF-1α对高糖诱导心肌线粒体损伤与细胞凋亡的影响

目的探讨盐酸右美托咪对高糖诱导的心肌线粒体损伤与细胞凋亡的影响及作用机制。方法用葡萄糖浓度为33 mmol/L的培养基培养H9C2细胞作为高糖组,正常培养基培养的细胞作为正常对照组;用1μmol/L的盐酸右美托咪定预处理H9C2细胞1 h后用葡萄糖浓度为33 mmol/L的培养基培养作为盐酸右美托咪定+高糖组;将con小干扰RNA(si-con)、HIF-1α小干扰RNA(si-HIF-1α)质粒分别转染至H9C2细胞中用1μmol/L的盐酸右美托咪定预处理H9C2细胞1 h后用葡萄糖浓度为33 mmol/L的培养基培养作为盐酸右美托咪定+si-con+高糖组、盐酸右美托咪定+si-HIF-1α+高糖组。四甲基偶氮唑盐比色法(MTT)检测细胞活力;乳酸脱氢酶(LDH)试剂盒检测LDH活性;蛋白质印迹(Western Blot)法检测裂解半胱氨酸天冬氨酸蛋白酶-3(Cleaved caspase-3)、B细胞淋巴瘤/白血病-2(Bcl-2)、Bcl-2相关X蛋白(Bax)、细胞色素c(Cyt-C)、缺氧诱导因子-1α(HIF-1α)蛋白表达水平;流式细胞术检测细胞凋亡;JC-1荧光标记法检测线粒体膜电位;2’,7’-二氯荧光黄双乙酸盐(DCFH-DA)荧光探针法检测活性氧(ROS)荧光强度;实时荧光定量PCR(RT-q PCR)检测细胞HIF-1αmRNA水平。结果与正常对照组相比,高糖组心肌细胞活力显著降低,LDH活性显著升高,Cleaved caspase-3、Bax表达水平显著升高,Bcl-2表达水平显著降低,细胞凋亡率显著升高,线粒体膜电位降低的细胞比率显著升高,Cyt-C蛋白表达水平显著升高,ROS荧光强度显著升高,HIF-1αmRNA和蛋白表达显著降低(P <0. 05)。与高糖组相比,盐酸右美托咪定+高糖组心肌细胞活力显著升高,LDH活性显著降低,Cleaved caspase-3、Bax表达水平显著降低,Bcl-2表达水平显著升高,细胞凋亡率显著降低,线粒体膜电位降低的细胞比率显著降低,Cyt-C蛋白表达水平显著降低,ROS荧光强度显著降低,HIF-1αmRNA和蛋白表达显著升高(P <0. 05)。与盐酸右美托咪定+si-con+高糖组相比,盐酸右美托咪定+si-HIF-1α+高糖组可逆转上述盐酸右美托咪定对高糖诱导的心肌细胞线粒体损伤与细胞凋亡的抑制作用。结论盐酸右美托咪定可促进心肌细胞存活,抑制LDH的漏出和ROS的产生,抑制细胞凋亡,稳定线粒体膜电位,减轻高糖对线粒体和细胞凋亡的影响,其机制可能与HIF-1α有关。     

Role of Reactive Oxygen Species during Low-Intensity Pulsed Ultrasound Application in MC-3 T3 E1 Pre-osteoblast Cell Culture

We evaluated the activation of mitogen-activated protein kinase (MAPK) activation through reactive oxygen species (ROS) by application of low-intensity ultrasound (LIPUS) to MC-3 T3 E1 pre-osteoblasts. The cells were subjected to one LIPUS application for either 10 or 20 min, and the control group was exposed to a sham transducer. For ROS inhibition, 10 μM diphenylene iodonium (DPI) was added to the cells an hour before LIPUS application. Samples were collected 1, 3, 6, 12 and 24 h after LIPUS application, and cells were evaluated for ROS generation, cell viability, gene expression and MAPK activation by immunoblot analyses. LIPUS caused a significant increase in ROS and cell viability in the non-DPI-treated group. Expression of RUNX2, OCN and OPN mRNA was higher in the LIPUS-treated groups at 1 h in both the DPI-treated and non-DPI-treated groups; RUNX2 and OCN mRNA levels increased at 6 h. ERK1/2 activation was increased in the LIPUS-treated groups. These results indicate that LIPUS activates MAPK by ROS generation in MC-3 T3 E1 pre-osteoblasts.