低剂量辐射抑瘤机制及联合化疗的研究

低剂量辐射具有广泛的应用前景，具有增强机体免疫功能、超敏感性等多种生物学效应，但是其具体的抑瘤的机制目前还不是很明确，现就低剂量辐射的抑瘤机制及联合化疗的研究和应用作一综述。     

Ad-EGFP-MDR1转染对小鼠骨髓细胞的影响

目的：通过研究Ad-EGFP-MDR1转染的骨髓细胞的细胞周期,凋亡相关蛋白表达和增殖分化有无改变,探讨基因修饰的骨髓细胞用于治疗的可行性。方法：用重组腺病毒Ad-EGFP-MDR1感染小鼠骨髓细胞,采用体外集落培养观察转染骨髓细胞与未转染细胞的增殖能力。采用免疫印迹检测两组细胞有无凋亡相关蛋白Bax、Caspase-9、p53的表达差异。使用流式细胞仪检测两组细胞的细胞周期有无差异。通过锥虫蓝拒染率判断细胞存活情况。结果：与对照组骨髓细胞比较,转染Ad-EGFP-MDR1的骨髓细胞体外集落形成差异无统计学意义。在转染的骨髓细胞和对照细胞中均未检测到凋亡相关基因Bax,Caspase-9,p53的表达,转染骨髓细胞与未转染对照骨髓细胞比较细胞周期差异无统计学意义（P>0.05）。锥虫蓝染色两组细胞的拒染率差异无统计学意义（P>0.05）。结论：骨髓细胞转染重组腺病毒Ad-EGFP-MDR1后其细胞凋亡相关基因,细胞增殖分化能力及细胞活性无明显变化,有利于进一步开展基因修饰骨髓细胞体内回输和体内安全性检测。     

NP方案联合不同间隔时间放疗对乳腺癌MCF-7细胞凋亡影响的研究

目的：研究NP方案[顺铂（DDP）＋盖诺（NVB）]联合不同间隔时间放射治疗对乳腺癌MCF-7细胞凋亡的影响。方法：采用流式细胞仪技术定性定量研究NP方案联合不同间隔时间放疗对人乳腺癌MCF-7细胞凋亡的影响,并观察NP方案作用后不同时间周期阻滞情况。结果：MCF-7细胞单独NP方案化疗组或放化联合作用组均可诱导细胞凋亡,随时间增加呈不可逆转增加,放化疗组凋亡指数明显高于单独化疗组,P〈0．05。NP化疗后不同间隔时间G2／M期阻滞比例不同。结论：化放不同间隔组凋亡指数随间隔时间延长而增加,间隔72h达到峰值。     

低剂量辐射抑瘤机制及联合化疗的研究

低剂量辐射具有广泛的应用前景,具有增强机体免疫功能、超敏感性等多种生物学效应,但是其具体的抑瘤的机制目前还不是很明确,现就低剂量辐射的抑瘤机制及联合化疗的研究和应用作一综述。     

低剂量辐射诱导小鼠脾脏细胞凋亡的适应性反应(英文)

Objective:The aim of the study was to investigate the adaptive response of low-dose radiation-induced apoptosis in mouse spleen cells and its related proteins control mechanism. Methods: Kunming male mice were randomly divided into sham-irradiation group, attacking dose irradiation group (D2) and low-dose radiation + attacking dose irradiation group (D1+D2). After a week the D1+D2 group was given low-dose radiation (doses were 25,50,75 and 100 mGy, dose rate of 12.5 mGy/min), after 6 hours the D2 group and ...     

逆转录病毒载体介导的双耐药基因转染小鼠骨髓细胞的实验研究

目的:将耐药基因以逆转录病毒为载体转染骨髓细胞以解决大剂量化疗所造成的骨髓抑制。方法:以逆转录病毒为载体,将双突变的二氢叶酸还原酶基因(DH-FR)和胞苷脱氨基酶基因(CD)转染小鼠骨髓细胞,观察该细胞耐MTX及Ara-C的CFU-GM生成情况;RT-PCR检测转基因的表达情况。结果:转基因的小鼠骨髓细胞有耐药克隆形成(14%);基因转染后小鼠骨髓细胞对MTX和Ara-C的耐受明显增加,P<0·005;RT-PCR显示转基因细胞有转染的基因条带。结论:双耐药基因通过逆转录病毒转染小鼠骨髓细胞并且获得表达,提高了骨髓细胞对MTX和Aar-C的耐药性。     

依马替尼对c-Kit+急性髓细胞白血病骨髓细胞凋亡的作用

目的观察依马替尼(STI571)体外对干细胞受体阳性(c-Kit )急性髓细胞白血病(AML)患者骨髓细胞凋亡的作用。方法用RT-PCR法检测c-Kit AML细胞抑凋亡基因SurvivinmRNA和促凋亡基因SmacmRNA的表达水平。结果抑凋亡基因SurvivinmRNA的表达水平下降,促凋亡基因SmacmRNA的表达水平上升。结论STI571能诱导c-Kit AML细胞促凋亡基因SmacmRNA上调,抑凋亡基因SurvivinmRNA的表达水平下降,发挥促凋亡作用。     

SOD对电离辐射诱导不同肿瘤细胞ROS水平和凋亡的影响

目的 研究ROS产生与电离辐射诱导细胞凋亡的关系，探讨SOD对：H22肝癌、Lewis肺癌、Hela宫颈癌细胞辐射敏感性的影响及机制。方法利用化学比色法测定肿瘤细胞中的ROS的水平，利用流式细胞仪测定肿瘤细胞凋亡。结果X线照射三种肿瘤细胞可通过产生ROS而诱发其凋亡，应用SOD在照射16h后对Lewins细胞的ROS水平无明显影响，而H22细胞和Hela细胞的ROS水平明显降低。SOD对：Hela宫颈癌细胞具有辐射保护作用，对Lewis肺癌细胞具有辐射增敏作用，对H22肝癌荷瘤小鼠肿瘤细胞辐射增敏性无明显影响。结论SOD通过改变肿瘤细胞的：ROS水平和参与其凋亡的分子生物学机制影响肿瘤细胞的辐射增敏性。     

低剂量X射线照射对K562细胞周期及凋亡影响的实验研究

目的 低剂量照射(low-dose radiation,LDR)可调节细胞信号传导,通过各种基因和蛋白的表达改变其分子生物学效应,由此产生了多样复杂且不同于大剂量照射的生物效应.用LDR及LDR联合大剂量照射(high-dose radiation,HDR)人红白血病细胞系K562(CML),探讨LDR对K562细胞周期进程及凋亡的影响.方法 按照K562细胞照射剂量不同分为4组,对照组(0 Gy)、LDR组(0.08、0.50 Gy)、HDR组(6 Gy)及LDR联合HDR组(0.08/6 Gy、0.50/6 Gy,LDR/HDR组).采用6 MVX射线照射,LDR与HDR间隔8 h;照射后按不同时间点收集细胞,流式细胞仪检测细胞周期变化与凋亡并分析细胞DNA倍体变化.结果 LDR后6h各组S期比例增加,对照组与LDR各组分别为55.38±2.22 vs 71.91±7.30、73.13±4.09(Z=-2.428,P=0.022;Z=-3.987,P＜0.001);12 h G2/M期细胞比例增加,出现G2/M阻滞,24 h达峰值,对照组与LDR各组分别为17.81±1.27 vs 26.61±7.82、29.02±2.76(Z=-3.684,P＜0.001;Z=-2.928,P＜0.001);48 h各组G0/G1细胞增多,72 h达峰值,对照组与LDR各组分别为27.07±1.19 vs 52.32±2.42、44.06±1.90(Z=-2.351,P=0.020;Z=-2.172,P=0.032).LDR/HDR后6hS期细胞比例增多,且G2/M期比例增加,即G2/M期阻滞,12 h达峰值并持续至24 h,12 h HDR组与LDR/HDR各组G2/M期分别为26.98±2.15 vs 56.27±1.57、69.31±2.51(Z=-2.564,P=0.021;Z=-7.759,P＜0.001).LDR后48 h凋亡率增加,96 h达峰值,对照组与LDR各组分别为1.82±0.12 vs3.21±0.20、6.28±0.30(Z=-3.959,P=0.003;Z=-9.705,P＜0.001);LDR/HDR照射后24 h凋亡率增加,120 h达峰值,HDR组与LDR/HDR各组分别为14.21±0.61 vs 17.38±0.92、27.91±1.07(Z=-2.986,P=0.027;Z=-6.973,P＜0.001).LDR后6h各组四倍体及八倍体细胞增加,且随照射剂量增大而增多,对照组与LDR各组八倍体细胞分别为2.41±0.15 vs 4.84±0.46、7.83±0.59,差异有统计学意义,H值分别为5.956和7.200,P值分别为0.025和0.001;LDR/HDR后6h各组四倍体及八倍体细胞增加,亦随LDR剂量增大而增多,HDR组与LDR/HDR各组八倍体细胞分别为6.49±1.05 vs 10.80±1.23、13.77±0.79,差异有统计学意义,H值分别为5.600和7.200,P值分别为0.05和0.001.结论 LDR能诱导K562凋亡,并能增强随后HDR对K562凋亡作用;LDR能诱导S/M期解偶联及G2/M期阻滞等细胞周期进程改变;LDR诱导K562凋亡可能与S/M期解偶联及G2/M期阻滞等改变细胞周期进程有关.     

γ线引起小鼠骨髓造血细胞凋亡的定量病理研究

迄今，骨髓辐射损伤后造血细胞凋亡的病理研究很少，其定量变化规律更未见报道。本文采用６０Ｃｏγ射线照射ＬＡＣＡ小鼠，于照射后６小时、１天及３天分批活杀，将右股骨骨髓经３％戊二醛及１％锇酸固定，制作半薄切片、超薄切片，并用ＣａｍｂｒｉｄｇｅＱｕａｎｔｉｍｅｔ９７０型图像分析仪，定量研究了辐射诱导的骨髓造血细胞凋亡的形态特征及其与照射剂量之间的关系。结果表明：对照组动物骨髓仅有少数造血细胞凋亡，而照射后６小时各剂量组动物骨髓内发生凋亡的造血细胞均明显增多，病理特点为：凋亡早期见核染色质浓缩、边移，呈半月形、环形或不规则形；中期见大小不等，形状不一的染色质团块周围有核膜包绕，核碎片形成；晚期见凋亡小体形成，可游离于细胞间或被邻近细胞吞噬。定量分析示各照射组动物凋亡的骨髓造血细胞的数密度和面密度均明显增加，与对照组比，其差异显著（Ｐ＜００１）。且剂量越大，凋亡细胞的数密度和面密度越大。最后，本文作者根据实验结果指出：凋亡是辐射介导的骨髓造血细胞死亡的主要死亡方式。本研究为急性放射病的诊治提供直接依据，亦可能对其他原因引起的细胞凋亡的研究具有参考意义。     

乳腺癌调强放疗与骨髓抑制的相关研究

目的 乳腺癌放疗时,患侧部分肋骨及胸骨均在照射野范围内,推测其受照的剂量-体积可能与骨髓增殖相关;加之放疗前多周期的化疗,骨髓增殖活性明显低下,放疗时部分患者因重度骨髓增殖抑制影响治疗的进程,进而影响总体疗效.分析乳腺癌放疗中骨髓抑制的相关因素,以期通过改进放疗靶区勾画及剂量限制,降低骨髓抑制发生率与严重程度,提高患者生活质量.方法 通过临床对照研究,收集徐州市肿瘤医院(40例)和徐州医学院附属医院(39例)2013-08-01-2014-08-26合计79例乳腺癌保留乳房术后患者,其中放疗中无骨髓抑制21例,1级骨髓抑制37例,2级骨髓抑制21例,采集每位患者的胸骨和肋骨受照剂量-体积(dose volume histograms,DVH)参数V5、V10、V20、V30、V50、D20、D40、D60、D80、D100、Dmin、Dmax和Dmean,分析各参数与骨髓增殖抑制之间的相关性.结果 肋骨参数比较,0级和2级之间比较,V5和Dmean差异有统计学意义,P值分别为0.038和0.027;1级和2级比较,V5(P=0.042)、V10(P=0.023)、V20(P=0.048)、D60(P=0.043)、D100(P=0.028)、Dmin(P=0.011)和Dmean(P=0.027)各参数之间差异均有统计学意义.胸骨参数比较,0级和1级除外V50和Dmax两个参数间差异无统计学意义,其余参数间差异均有统计学意义,P＜0.05.1级与2级V30和V40之间差异有统计学意义,P值分别为0.037和0.024.结论 肋骨、胸骨受照的DVH相关参数对骨髓增殖活性的影响有意义;与肋骨相比,胸骨对射线更为敏感.     

辐射损伤骨髓造血细胞的DNA含量及AgNOR定量分析

LACA小鼠经60 Coγ射线全身一次照射 ,对骨髓组织切片和骨髓细胞悬液涂片 ,分别采用改良的Feulgen染色及核仁组成区相关蛋白 (AgNOR)染色 ,利用图像分析技术对在不同照射剂量、不同时间点的造血细胞核内DNA及AgNOR含量进行了定量分析。结果显示各照射组在2 5 - 7 0Gy剂量、 6h～ 4周范围内 ,骨髓均出现明显损伤及损伤后重建现象 ,且剂量越大 ,损伤越重 ,恢复亦较慢 ;其中以 5 5Gy组骨髓细胞核内DNA和AgNOR含量变化最为典型 ,在损伤早期进行性减少 ,而在恢复期出现持续性增加 ,直至恢复正常。结果提示 :骨髓造血细胞核内AgNOR及DNA含量的变化可以作为反映骨髓辐射损伤与修复程度的定量指标。     

Temporal response of murine bone marrow to local hyperthermia

Hemopoietic insufficiency resulting from single or multiple agents used in cancer therapy often becomes a dose limiting toxicity. Since hyperthermia is finding increased clinical application with whole body or extended field treatments for a variety of neoplastic diseases, we wished to examine its potential effects on normal bone marrow. One hind leg of anesthetized mice received up to one hour water bath heating to 41–44°C. With temperatures of 42°C or greater, there was a dose and heating time dependent initial depletion of both CFU-S and CFU-GM that was not appreciably affected by migration of stem cells to or from the heated field. CFU-S and CFU-GM returned to normal values by 2 to 3 weeks after 1 hour heating to 42°C. However, at 43° or 44°C for 1 hour, CFU-GM only transiently returned to normal, falling to 55% and 33% of control respectively by 3 months. Total nucleated cell counts reflected a similar dose and time dependent depletion and repopulation. Finally, bone marrow differential counts from heated marrow suggested that equivalent repopulation along each differentiation pathway occurred during recovery after thermal injury. These data indicate that temperatures greater than 42°C for one hour caused a long term delay in hemopoietic repopulation, which may contribute to a significant toxicity when hyperthermia is used in combination with chemotherapeutic drugs or local irradiation incorporating the same treatment field.     

46. Micronuclei induced by chronical treatment of SO2 inhalation in mouse bone marrow cells

In the chronical experiment of treating with sulfur dioxide(SO2) inhalation, Micronuclei(MN) frequencies in the polychromatophilic erythroblasts(PCE) of mouse bone marrow and the frequencies of cells with MN were significantly increased in dose-dependent manner. There is a significant difference between the male and the female animals. The results also showed that SO2 inhibited urethone-induced MN formation, it is a antagonistic joint action to Urethone. These results furtherly confirm that SO2 inhalation is a clastogenetic and genotoxic agent to mammalian cells, and the combination roles of SO2 and other mutagens are complexity.     

Increased expression of CX43 on stromal cells promotes leukemia apoptosis

Connexin 43 (Cx43) induced apoptosis has been reported in solid tumors, but the effect of Cx43 expressed by bone marrow stromal cells (BMSC) in leukemia has not been fully investigated. Manipulating Cx43 expression could be a potential therapeutic strategy for leukemia. Here, we investigate the effect of Cx43 expressed by BMSCs (human Umbilical Cord Stem Cells over-expressed CX43, Cx43-hUCSC) on leukemia cells. When co-cultured with Cx43-hUCSC, leukemia cells show significant lower growth rate with increasing apoptosis activity, and more leukemia cells enter S phase. Functional assays of fluorescence recovery after photo bleaching (FRAP) showed improved gap junctional intercellular communication (GJIC) on leukemia cells when co-cultured with Cx43-hUCSC (p < 0.01). In a mouse minimal disease model, the mean survival time and mortality rate were significantly improved in mice transplanted with Cx43-hUCSC. Our results indicate that Cx43 expressed by BMSC induces apoptosis on leukemia cells. Small molecules or other pharmaceutical approaches for modulating Cx43 expression in BMSCs could be used for delaying relapse of leukemia.     

Further evidence for the existence of ‘homing’ receptors on murine leukemia cells which mediate adherence to normal bone marrow stromal cells

A significant proportion of ¹³¹IUDR-labelled cells from murine leukemia cell lines L1210 and P388, but not the L5178Y lymphoma cell line, are retained in the bone marrow (B.M.) following i.v. injection into syngeneic mice. Following this, L1210 and P388 cells grow and rapidly replace the normal hematopoietic cells of the B.M. L1210 and P388 cells, but not several lymphoma cell lines, also bind avidly to monolayers of B.M. stromal cells (Dexter cultures) and soon overgrow the cultures following rapid cell proliferation. P388 cells bound equally well to confluent monolayers of B.M., whole mouse embryo and newborn mouse kidney while L1210 cells bound well to B.M. and whole mouse embryo but showed little binding to newborn kidney monolayers. The accumulation of the two leukemia cell lines in the B.M. was constant and indistinguishable over a 48-h period. In contrast, in both spleen and liver the number of L1210 cells decreased during the same period while P388 cells were retained at a constant level. Generally there was a lack of correlation of B.M. metastasis of a cell line and its metastasis to other organs although P388 cells, but not L1210 cells, demonstrated a tremendous capacity for metastatic growth in both spleen and liver.

Normal B.M. cells were fused with the syngeneic SP2/0 murine myeloma fusor line and 10 hybridomas plus the SP2/0 parent were tested for in-vitro adherence to B.M. monolayers and in-vivo metastatic behavior. The same 3 (out of 10) hybridomas showed a high level of adherence to B.M. monolayers, high levels of retention of cells in the B.M. following i.v. injection, and rapid growth and takeover of the normal B.M. In marked contrast, neither the SP2/0 parent nor the remaining 7 hybridomas show significant adherence, B.M. retention or growth in the B.M. A distinct lack of correlation of B.M. vs liver or spleen metastasis was once again noted for the hybridomas although all of the hybridomas showed much less metastatic growth in the liver than the SP2/0 parent. Seven out of 10 hybridomas also showed less metastatic growth in the spleen including all 3 of the hybridomas which showed preferential growth in the B.M.

Our data are consistent with the existence of cell surface ‘homing’ receptors on leukemia cells for normal B.M. stromal cells which function to retain the leukemia cells in the B.M. Such receptors might serve as functional markers for cell differentiation and leukemia classification.     

黄芪甲甙调控Nrf-2表达对骨髓间充质细胞凋亡机制的研究

目的:研究黄芪甲甙是否通过Nrf2-Keap1/ARE信号通路发挥抗氧化作用来抑制骨髓间充质细胞的凋亡。方法:培养Wistar大鼠BMSCs,设立正常组和黄芪甲甙干预组。采用实时荧光定量RT-PCR方法测定Nrf-2以及GSH的基因表达,Westem blot检测其蛋白表达。结果:TUNEL和流式结果表明黄芪甲甙有抑制BMSCs凋亡的作用,50μg/L组作用较强,与ADR组比较差异有显著性。增加浓度抑制凋亡的作用增加不明显,ADR+AST(100μg/L、50μg/L)两组,细胞凋亡率差异无显著性。黄芪甲甙作用后明显降低Nrf2 mRNA的表达,与ADR模型组比较差异有统计学意义。各组分别与阿霉素模型组比较差异均有统计学意义。结论:黄芪甲甙可以通过Nrf-2表达调控细胞凋亡,可能是其抑制神经细胞氧化应激、发挥保护作用的机制之一。