目的 构建吉西他滨耐药乳腺癌细胞4T1耐药株并建立裸鼠乳腺癌肝转移模型。方法 采用低浓度加量持续诱导法,诱导吉西他滨耐药乳腺癌细胞4T1耐药株,命名为4T1/Gem;CCK-8法测定4T1与4T1/Gem细胞的增殖抑制率,计算耐药指数; Western blot法检测细胞P-gp蛋白表达;B超引导下注射4T1/Gem细胞悬液诱导裸鼠肝脏成瘤;HE染色观察肿瘤组织病理情况,免疫组化法检测瘤组织ER、PR、HER2、Ki-67和P-gp蛋白的表达。结果 经过14个月的诱导成功建立4T1/Gem细胞株,可在含40 μg/mL的Gem培养液中稳定生长。4T1/Gem细胞耐药指数为4T1细胞的788.547倍。与亲代相比,4T1/Gem处于G1期和G2期的细胞增加,S期细胞减少;上调P-gp蛋白的表达。4T1/Gem细胞成功建立裸鼠乳腺癌肝转移模型,瘤组织中ER、PR、HER2蛋白阴性表达,Ki-67阳性10％和P-gp蛋白阳性表达。结论 成功构建吉西他滨耐药乳腺癌细胞4T1耐药株并建立裸鼠乳腺癌肝转移模型,为开发治疗乳腺癌肝转移化疗耐药的药物提供实验基础。
Objective To construct a gemcitabine-resistant variant of the breast cancer cell line (4T1/Gem) and establish a nude mouse model of breast cancer with hepatic metastatic. Methods A gemcitabine-resistant variant of the breast cancer 4T1 cell line was induced by gradually increasing the concentration of gemcitabine; this variant is referred to in this study as 4T1/Gem. The proliferation suppression rates of 4T1 and 4T1/Gem cells were determined by using the CCK-8 essay to evaluate the drug resistance indices of the cell lines. Western blot analysis was used to detect P-gp protein expression. Under ultrasonography, a 4T1/Gem cell suspension was injected into nude mice to induce liver tumors. H&E staining was used to observe tumor pathology, and immunohistochemistry was used to detect the expression of ER, PR, HER-2, Ki-67, and P-gp. Results After 14 months of induction, a 4T1/Gem cell line is established successfully. The cell line can grow stably in culture liquid containing 40 μg/ml gemcitabine. The drug resistance index of 4T1/Gem is 788.547. Compared with the 4T1 cell line, the 4T1/Gem cell line can upregulate P-gp protein expression and successfully establish a nude mouse model of breast cancer with hepatic metastatic. ER, PR, and HER-2 proteins exhibit negative expression in the tumor tissue. The positive expression of P-gp and 10% of Ki-67 proteins is also observed. Conclusion This study successfully constructs a gemcitabine-resistant variant of the breast cancer cell line (4T1/Gem)and establishes a nude mouse model of breast cancer with hepatic metastatic, thereby providing an experimental basis for developing and treating a drug-resistant variant of breast cancer.
目的 探讨胰腺神经鞘瘤的临床特点和诊治方法。方法 总结并回顾性分析我院肝胆外科收治的胰腺神经鞘瘤患者1例及文献报道的71例患者临床资料。结果 共计72例胰腺神经鞘瘤患者纳入总结和分析。患者平均年龄54岁(范围17～89岁),其中女性40例(56%)。临床表现包括上腹痛、体重减轻,或体检偶然发现胰腺肿物。肿瘤平均大小6.1 cm(1～20 cm)。肿瘤位于胰头部29例(40%)、胰体/尾部32例(44%),沟突部6例(8%)。肿瘤表现为实性肿物27例(38%)、囊性28例(39%)、囊实性10例(14%)。2例通过术前超声内镜下穿刺活检病理确诊,其余均为手术后标本病理诊断证实。手术治疗行胰十二指肠切除术23例、局部剜除术16例、胰体尾切除术15例、胰腺中段切除1例。5例 (7%) 患者术后病理为恶性神经鞘瘤,恶性组肿瘤大小明显大于良性组[(13.8±6.2)cm vs (5.6±4.1)cm,P=0.0004)]。手术切除患者术后随访3～65月,均无肿瘤复发、转移及患者死亡。结论 胰腺神经鞘瘤临床表现缺少特异性,术前诊断困难,肿瘤大小与良恶性具有明显相关性,手术治疗可取得良好效果。
Objective To analyze clinical presentation, diagnosis, treatment options, and outcome of pancreatic schwannoma. Methods A retrospective study of clinical data of a case in our hospital and 71 cases reported in literature with pancreatic schwannoma. Results 72 cases were analysed. The mean age was 54 years (range 17-89 years), with 56 % of patients being female. Mean tumor size was 6.1 cm (range 1-20 cm). Tumor location was the head (29 cases), body and tail (32 cases), and uncinate process (6 cases). 27 cases exhibited solid tumors and 28 cases exhibited cystic tumors. Treatment included pancreaticoduodenectomy (23 cases), distal pancreatectomy (15 cases), enucleation (16 cases). 5 cases (7%) were malignant schwannoma. Tumor size of malignant group was significant larger than benign group (13.8±6.2 cm vs 5.6±4.1 cm,P=0.0004). There was no local recurrence metastasis,or death at the follow-up after operation (range 3-65 months). Conclusion The clinical manifestations of pancreatic schwannoma are lack of specificity and preoperative diagnosis remains difficulty. The tumor size was significantly related to classification of malignant or benign. Pancreatic schwannoma has satisfactory prognosis with surgical treatment.
目的 探讨生理情况下高龄肝脏与低龄肝脏的区别,寻找可以区分高龄肝脏与低龄肝脏的指标。方法 取6周龄SD大鼠和9月龄以上退役SD大鼠各5只,采用超声弹性成像检测肝脏硬度、全自动生化检测仪检测血清学指标、H&E染色观察肝脏形态结构、Sirius Red染色及Masson染色检测胶原纤维的沉积、免疫组化SP法检测TGF-β1、p16INK4a、SMP-30蛋白的表达。结果 高龄组和低龄组之间血清学指标、胶原纤维沉积及TGF-β蛋白、p16INK4a蛋白的表达无差异;超声弹性成像检测低龄组Vs值为(1.21±0.09)m/s,高龄组为(1.32±0.05)m/s(P=0.033);SMP-30蛋白低龄组IOD值为138244.988±51286.257,高龄组为116240.170±35017.936(P=0.007)。结论 高龄大鼠与低龄大鼠肝脏的硬度及SMP-30蛋白的表达存在差异,随着年龄的增加肝脏硬度增大,SMP-30蛋白表达下降。肝脏硬度与SMP-30蛋白可作为区分高龄肝脏与低龄肝脏的指标。
Objective To investigate the differences between elderly and younger liver. Methods In accordance with the age of the SD rats into two groups: younger group (Group Y, 6 weeks, n=5) and elderly group (Group O, 40 weeks or more, n=5). Data were compared by using ultrasound elasticity imaging to detect liver stiffness, automatic biochemical detector to gauge serum indexes, H&E staining to observe the liver morphological structure, Sirius Red staining and Masson staining to assay the collagen fibers deposition, Immunohistochemistry to identify the expression of TGF-β1, p16INK4a and SMP-30 protein. Results Serum indexes, collagen deposition, TGF-β1 and p16INK4a protein expression were no statistically significant difference between two groups. The Vs value was (1.21±0.09) m/s in Group Y and (1.32±0.05) m/s in Group O (P=0.033). and the IOD value of SMP-30 protein between Group Y and Group O were 138244.988±51286.257 and 116240.170±35017.936 (P=0.007). Conclusion The degree of liver stiffnessnd and SMP-30 protein in elderly and younger liver are different.Increased the degree of liver stiffness and decreased the expression of SMP-30 protein in the elderly SD rats. Liver stiffness and SMP-30 protein could be used as indicators to distinguish between elderly and younger liver.
目的 动态观察乳腺癌患者辅助内分泌治疗5年后的血脂及肝功能水平的变化,探求辅助内分泌治疗与高脂血症及脂肪肝发病率的关系。方法 56例乳腺癌患者实行辅助内分泌治疗,术后随访5年动态抽血测定其总胆固醇(TC)、甘油三酯(TG)、低密度脂蛋白胆固醇(LDL)、高密度脂蛋白胆固醇(HDL)及谷草转氨酶(AST)、谷丙转氨酶(ALT)、直接胆红素(DBIL)、总胆红素(TBIL)等参数的变化,B超监测其肝脏变化。结果 经过2年内分泌治疗TG由(1.203±0.723)mmol/L上升至(1.701±1.271)mmol/L,5年内分泌治疗后TG降至(1.389±0.706)mmol/L。经过2年内分泌治疗LDL由(2.497±0.990)mmol/L上升至(2.950±0.984)mmol/L,5年内分泌治疗后LDL为(2.867±0.886)mmol/L。结论 辅助内分泌治疗2年会导致其TG和LDL的升高,5年随访仅发现LDL升高,辅助内分泌治疗会增加乳腺癌患者诱发心血管疾病的风险。
目的 初步探究胆管结扎诱导的梗阻性胆汁淤积对大鼠肝细胞的影响。方法 10只Lewis大鼠随机分为对照组和胆汁淤积组,每组各5只,胆汁淤积组采用胆管结扎2周诱导梗阻性胆汁淤积大鼠模型。苏木精-伊红染色和苯胺蓝染色比较组织病理变化,使用生化分析比较两组小鼠肝功能情况。采用改良的两步胶原酶灌注分离原代肝细胞,通过RT-qPCR检测两组小鼠肝细胞标志基因、细胞增殖标志基因以及胆管细胞标志基因的表达情况。结果 与对照组相比,胆汁淤积组肝脏表现为明显的肝组织紊乱和纤维胶原蛋白沉积以及肝功能的损害。胆汁淤积组较对照组的原代肝细胞更高表达细胞增殖标志基因：细胞增殖标志物（Ki67）基因,叉头盒M1蛋白（Foxm1）基因,增殖细胞核抗原（Pcna）基因和肝细胞生长因子（HGF）基因（P<0.05）;胆汁淤积组的原代肝细胞表达更低水平的肝细胞标志基因：白蛋白（Alb）基因,多药耐药相关蛋白2（Mrp2）基因,胆盐输出泵（Bsep）基因和肝细胞连环蛋白1（Catenin1）基因（P<0.05）,同时表达更高水平的胆管细胞标志基因：细胞角蛋白7（Ck7）基因,细胞角蛋白 19（Ck19）基因,胆管细胞多药耐药性蛋白1（Mdr1）基因和胆管细胞囊性纤维化跨膜传导调节因子（Cftr）基因（P<0.05）以及肝祖细胞标志基因：上皮细胞黏附分子（Epcam）基因和Y染色体性别决定区-盒转录因子9（Sox9）基因（P<0.05）。结论 胆汁淤积可诱导肝细胞向胆管细胞特性转化的可塑性。
Objective To explore the effect of bile duct ligation induced obstructive cholestasis on rat hepatocytes. Methods Ten Lewis rats were randomly divided into control group and cholestasis group, and the cholestasis was induced by bile duct ligation for 2 weeks. The histopathological changes were compared by H&E and aniline blue staining and the liver function was compared by biochemical analysis. Primary hepatocytes were isolated by modified two-step collagenase perfusion, and the expressions of hepatocyte marker genes, cell proliferation marker genes and cholangiocyte marker genes were detected by RT-qPCR. Results Compared with the control group,the liver of the cholestatic group showed obvious disordered histopathology, deposition of fibrous collagen and impaired liver function. Compared with the control group, the primary hepatocytes in the cholestasis group expressed higher cell proliferation-related genes（Ki67,Foxm1,Pcna and HGF）（P<0. 05）. Primary hepatocytes in the cholestasis group expressed lower levels of hepatocyte marker genes（Alb,Mrp2,Bsep and Catenin1）（P<0. 05）,and higher levels of cholangiocyte marker genes（Ck7,Ck19,Mdr1 and Cftr）（P<0. 05）and higher levels of the hepatic progenitor cell marker genes（Epcam and Sox9）（P<0. 05）. Conclusions Cholestasis induces rat hepatocyte plasticity in the transformation into bile duct properties.
