Morphological changes in hepatic vascular endothelium after carbon dioxide pneumoperitoneum in a murine model

Background: Liver metastasis is an important prognostic factor in advanced colorectal cancer. Several studies have demonstrated that carbon dioxide (CO₂) pneumoperitonem enhances liver metastasis in an animal model. In the present study, we used scanning electron microscopy (SEM) to investigate morphological changes in hepatic vascular endothelium after CO₂ pneumoperitoneum in a murine model.Methods: Thirty-three male BALB/c mice were randomized to undergo pneumoperitoneum (CO₂, air, or helium ), open laparotomy, and anesthesia alone. After each procedure, the animals’ livers were excised at days 0, 1, and 3 and examined by SEM.Results: In the CO₂ pneumoperitoneum group, we observed rough surface and derangement of the hepatic vascular endothelial cells and intercellular clefts on day 1. In the other groups, no major morphologic changes were observed at any time.Conclusions: Hepatic vascular endothelium changes after CO₂ pneumoperitoneum. Such characteristic changes may play an important role in establishing liver metastasis after CO₂ pneumoperitoneum.     

Hyaluronic acid secretion during carbon dioxide pneumoperitoneum and its association with port-site metastasis in a murine model

Background: The mechanism of port-site metastasis after laparoscopic cancer surgery is unclear. This study aimed to determine whether carbon dioxide (CO2) pneumoperitoneum caused an increase in hyaluronic acid, which is secreted from mesothelial cells of the peritoneal cavity, and to assess the risk for port-site metastasis using a murine pneumoperitoneal model. Methods: Sandwich-binding protein assay was used to measure the concentration of hyaluronic acid in the peritoneal cavity at 6, 12, 18, 24, 48, and 72 h after CO2 pneumoperitoneum or laparotomy for 30 min. The concentrations of hyaluronic acid during pneumoperitoneum were compared among different gases (CO2, helium, air), intervals (5, 30, 60 min), and pressures (0-2, 4-6, 8-10 mmHg). To investigate the effects of exogenous hyaluronic acid, the development of port-site metastasis was examined using mouse adenocarcinoma cell-line colon 26 cells. Results: The intraperitoneal concentration of hyaluronic acid after CO2 pneumoperitoneum had increased already at 6 h, had reached the maximum level at 24 h, and had begun to decrease at 72 h. The concentration of hyaluronic acid at 24 h and 48 h in the CO2 pneumoperitoneum group was higher than that in the laparotomy group. This increase in hyaluronic acid also was found during helium and air pneumoperitoneum, and the concentration of hyaluronic acid in the peritoneal cavity was at its maximum when CO2 pneumoperitoneum lasted 30 min at 4 to 6 mmHg. The frequency of port-site metastasis was the highest when hyaluronic acid was injected during CO2 pneumoperitoneum (100%). Conclusions: In a murine model, the intraperitoneal concentration of hyaluronic acid was significantly increased after CO2 pneumoperitoneum, and the increase was more evident than that after laparotomy. Increased hyaluronic acid during pneumoperitoneum may be associated with port-site metastasis after laparoscopic cancer surgery.     

Port-site metastasis after CO<Subscript>2</Subscript> pneumoperitoneum

Background Port-site metastasis is a continuing problem in laparoscopic cancer surgery. To clarify the role of adhesion molecules in the development of port-site metastasis, particularly with regard to prevention, we performed experiments in which port-site metastasis was inhibited using antibodies against extracellular matrix proteins or the active Arg–Gly–Asp (RGD) peptide after CO₂ pneumoperitoneum in a murine model.Methods We examined the development of port-site metastasis under the following conditions: (1) CO₂ pneumoperitoneum with or without hyaluronic acid and anti-integrin or anti-CD44 antibody and (2) CO₂ pneumoperitoneum and a RGD peptide or pseudo-RGD sequence peptide (FC-336). BALB/c mice (n = 130) were injected with 5 × 10⁵ human gastric cancer cells (MKN45) and either antibody or peptide, treated with CO₂ pneumoperitoneum, and injected intraperitoneally with antibody or peptide for 5 days. Three weeks after CO₂ pneumoperitoneum, the frequency and weight of port-site metastatic tumors were determined.Results Anti-integrin antibody significantly decreased the weight of port-site metastatic tumors without hyaluronic acid (control vs anti-integrin: 8.2 ± 7.1 vs 3.6 ± 4.5 mg; p < 0.05) but not the frequency of port-site metastases. With hyaluronic acid, the frequency of port-site metastasis and the weight of port-site metastatic tumors were significantly decreased both by anti-integrin and by anti-CD44 antibody (control vs anti-integrin and anti-CD44; 95% and 8.5 ± 7.2 mg vs 50% and 3.1 ± 4.3 mg and 55% and 3.3 ± 5.1 mg, respectively; p < 0.05). RGD peptide and FC-336 also inhibited port-site metastasis in a dose-dependent manner.Conclusion Cell adhesion molecules integrin and CD44 play an important role in the development of port-site metastasis after laparoscopic cancer surgery. Intraperitoneal injection of RGD peptide or pseudo-RGD sequence peptide (FC-336) can prevent port-site metastasis.     

Enhancement of port site metastasis by hyaluronic acid under CO2 pneumoperitoneum in a murine model

Background

The mechanism underlying the development and progression of port site metastasis after laparoscopic surgery for cancer is still not understood. Hyaluronic acid secreted from mesothelial cells is thought to be a key factor that causes adhesion between cancer cells and mesothelial cells. Using a murine model of carbon dioxide (CO₂) pneumoperitoneum, we evaluated the effect of exogenous hyaluronic acid on port site metastasis.

Methods

BALB/c mice were injected with 5×10⁶ human gastric carcinoma (MKN45) cells and divided into four groups treated with or without hyaluronic acid and with or without pneumoperitoneum. Three weeks later, the frequency and weight of port site metastases were determined. The effects of hyaluronic acid on tumorigenicity and tumor with MKN45 cells.

Results

Port site metastasis occurred significantly less frequently in the pneumoperitoneum-only group than in the pneumoperitoneum-with-hyaluronic-acid group (75% vs 100%, p<0.05). The port site metastatic tumor weighed significantly less in the control group (anesthesia only) than in the hyaluronic acid group (89±17 vs 288±35mg, p<0.05); it also weighed less in the pneumoperitoneum-only group than in the pneumoperitoneum-with-hyaluronic-acid group (87±24 vs 298±51 mg, p<0.05). The frequency and weight of tumors in the subcutaneous tissue were not significantly different between groups with or without hyaluronic acid injection (95% vs 90%, 331±128 vs 322±115 mg).

Conclusions

Under CO₂ pneumoperitoneum, exogenous hyaluronic acid increased the frequency and weight of port site metastasis in a murine model. Hyaluronic acid secreted from mesothelial cells may be associated with the formation of port site metastasis after laparoscopic surgery for cancer under pneumoperitoneum.     

Microcirculation and excretory function of the liver under conditions of carbon dioxide pneumoperitoneum

Background: To date, the effects of increased abdominal pressure, as given during carbon dioxide (CO₂) pneumoperitoneum, on hepatic microcirculation and biliary excretion are unknown.Methods: Using a custom-made peritoneal cavity chamber, we performed intravital microscopy of the left liver lobe under conditions of CO₂ pneumoperitoneum in a rat model. In addition, biliary excretion was assessed.Results: The establishment of a CO₂ pneumoperitoneum of 4 or 8 mmHg resulted in sinusoidal perfusion failure that was more pronounced in the periportal regions than in the midzonal and pericentral regions of the liver acinus. Biliary excretion was considerably reduced at an intraabdominal pressure of 8 mmHg. Leukocyte–endothelial cell interactions increased significantly in both hepatic sinusoids and postsinusoidal venules.Conclusion: Alterations in hepatic microcirculation and liver function must be taken into consideration in any kind of laparoscopic surgery and may be of particular clinical relevance in patients with liver pathology.     

肝移植术后移植肝组织P-选择素、ICAM-1基因mRNA表达的实验研究

目的：研究移植肝组织P-选择素、ICAM-1基因mRNA表达变化以及供肝交感神经、枯否细胞对mRNA表达的影响。方法：使用氯化钆抑制供肝枯否细胞、六甲胺阻断供肝交感神经，观察肝移植术后4，8，16，24h移植肝P-选择素、ICAM-1 mRNA表达的变化。结果：肝移植术后肝组织P-选择素、ICAM-1基因mRNA表达增高，阻断供肝交感神经和抑制供肝枯否细胞，可下调P-选择素、ICAM-1的mRNA表达。结论：阻断供肝交感神经和抑制供肝枯否细胞能明显下调肝组织P-选择素、ICAM-1基因表达，减轻移植肝的缺血再灌注损伤。     

三氧化二砷对CO2气腹下小鼠肿瘤腹壁戳口种植影响的实验研究

目的 研究腹腔内注射三氧化二砷(arsenic trioxide,As2O3)对小鼠CO2气腹下肝癌H22转移的影响. 方法 昆明鼠40只(清洁级),中腹部穿刺置入1 mm套管针,自套管针注入1×106肿瘤细胞后,建立CO2气腹,压力8 mm Hg,时间30 min.术后随机分4组,每组10只,分别腹腔内注入生理盐水,1 ml;As2O3(2 mg/kg),1 ml;As2O3(4 mg/kg),1 ml;As2O3(4mg/kg)+肝素(10 U/ml),共1 ml.气腹后第3、7天测量肿瘤黏附因子(CD44)、血管内皮生长因子(vascular endothelial growth factor,VEGF)的变化;比较各组生存状态、腹围、体重变化及转移瘤直径.结果 气腹后第3、7天,与对照组相比,各As2O3组CD44、VEGF表达均明显降低(P＜0.05).2个高剂量组的气腹后第3天VEGF、第7天CD44比低剂量组降低明显(P＜0.05).4组戳口种植率分别为9/10、8/10、7/10、6/10,差异无显著性(x2=2.667,P=0.446). 结论 As2O3对CO2气腹腹腔镜肿瘤生长转移有抑制作用.     

Influence of carbon dioxide pneumoperitoneum environment on adhesion and metastasis of a human ovarian cancer cell line

Background  This study aimed to explore the adhesion and metastasis capability of the human ovarian cancer cell line SKOV₃ after exposure to a simulated laparoscopic carbon dioxide (CO₂) pneumoperitoneum environment and the related mechanism. Methods  SKOV₃ was subjected to a simulated laparoscopic CO₂ pneumoperitoneum environment at various CO₂ pressures (8–12 mmHg) and exposure times (1–3 h). Cell adhesive capacity was determined by a mechanical method. Real-time fluorescent quantitative polymerase chain reaction (FQ-PCR) and immunocytochemical staining were used to examine the mRNA and protein expressions of heparanase (HPSE) and vascular endothelial growth factor C (VEGF-C) in SKOV₃. Cells cultured in a standard environment were used as the control. Results  The increase of SKOV₃ cell adhesion capability was associated with CO₂ pressure and exposure time. A significantly higher adhesion capability was observed in the group with exposure to 10 mmHg for 2 h over a 48 h period, as compared with the control groups (p < 0.05). The expressions of HPSE and VEGF-C in SKOV₃, which are closely related to metastasis capability, also increased. Significantly higher expressions were observed in the group with exposure to 10 mmHg for 3 h over a 48 h period, as compared with the control groups (p < 0.01). Conclusions  The adhesion and metastasis capacity of SKOV₃ increased with overexpression of HPSE and VEGF-C and were positively related to CO₂ pressure, exposure, and culture time.     

Liver histology alterations during carbon dioxide pneumoperitoneum in a porcine model

Background This study aimed to investigate the time course changes in liver histology during carbon dioxide (CO₂) pneumoperitoneum in a large animal model. Methods For this study, 14 white pigs were anesthetized. Liver biopsies performed 0, 1, and 2 h after establishment of CO₂ pneumoperitoneum (at 12 mmHg) and after peritoneal desufflation were sent for histologic examination. Heart rate, mean blood pressure, hepatic artery flow, portal vein flow, and aortic flow were recorded in 10-min increments. Three animals served as control subjects. Results A statistically significant time course increase was observed in portal inflammation, intralobular inflammation, edema, sinusoidal dilation, sinusoidal hyperemia, centrilobular dilation, centrilobular hyperemia, pericentrilobular ischemia, and focal lytic necrosis scores. There were no significant changes in the control group. This eliminated an effect of anesthesia only. The portal vein flow increased as much as 21%, and the hepatic artery flow decreased as much as 31% of baseline, but these differences did not attain statistical significance. Aortic flow remained relatively stable. Conclusion Histomorphologic changes occurred, indicating liver tissue injury during CO₂ pneumoperitoneum at an intraabdominal pressure of 12 mmHg in the porcine model. Portal vein flow increased, and hepatic artery flow decreased, whereas aortic flow remained relatively unaffected in this experiment. Presented in part orally at the 10th World Congress of Endoscopic Surgery, Berlin, September 2006     

ICAM-1mRNA在大鼠缺血/再灌注肝脏组织中的表达

目的探讨肝脏缺血/再灌注损伤过程中,肝脏组织中ICAM-1mRNA的表达规律及其意义。方法应用RT-PCR技术,观察缺血时间分别为15min、30min及45min的三组大鼠肝脏于再灌注60min时ICAM-1mRNA的表达情况。结果三组肝脏缺血前及缺血末组织内仅有少量ICAM-1mRNA表达于肝细胞,但于再灌注60min时,ICAM-1mRNA表达程度则显著增强,且缺血时间越长的肝脏,其表达强度越高。结论肝脏的缺血能明显诱导再灌注期间肝细胞表达ICAM-1mRNA,增强肝窦内皮细胞的粘附力,进而引发一系列病理生理改变。     

裸鼠肝癌转移模型组织中细胞间粘附分子1的表达

研究细胞间粘附分子１（ＩＣＡＭ－１）在肝癌转移过程中的作用和作为预测肝癌转移标志物的价值。方法以裸鼠人肝癌转移模型为材料,以ｄｏｔｉｍｍｕｎｏ－ｂｌｏｔ的方法测定不同生长时间肝癌组织中ＩＣＡＭ－１的表达。结果ＩＣＡＭ－１在肝癌组织中的表达与肿瘤的生长时间呈明显的正相关（ｒ＝０．８８,Ｐ＜０．０１）,与肿瘤大小呈正相关（ｒ＝０．０５,Ｐ＜０．０５）,在肝癌转移发生后ＩＣＡＭ－１表达突然升高,以后维持在较高水平,肿瘤转移组高于无转移组（Ｐ＜０．０１）,多脏器转移组高于单脏器转移组（Ｐ＜０．０５）。结论组织中ＩＣＡＭ－１的表达可以反映裸鼠人肝癌的生长转移状态,可以作为预测其转移的标志物。     

Liver metastasis following pneumoperitoneum with different gases in a mouse model

apd: 20 October 2000     

CO2气腹对肝硬化兔肝脏血流的影响

目的:探讨CO2 气腹对肝硬化兔肝脏血流循环及肝功能的影响。方法:制备肝硬化兔模型,于不同的CO2 气腹,用彩色多普勒超声腹腔内检测门静脉、肝动脉和背主动脉的血流速度、管内径和肝动脉阻力指数,监测心率,计算血流量、门静脉淤血指数。检测气腹前后兔肝功能。结果:腹内压升高,肝硬化组和对照组兔背主动脉血流、心率改变差异无显著性;门静脉血流降低,肝硬化组于10mmHg气腹压下减少45. 5% (P<0. 05),同时肝动脉血流增加,肝脏总体循环血流减少,肝功能受损,肝硬化兔改变稍明显,但两组差异无显著性。系统血供(背主动脉血流)与肝脏血流无关联(P>0. 05),门静脉血流与腹内压有相关性(P<0. 05),肝动脉血流与动脉阻力指数呈负相关(P<0. 05)。结论:CO2 气腹对肝硬化兔肝脏血流影响较大,门静脉血流显著降低且肝动脉缓冲效应减弱;肝硬化兔肝功能更易受损。尽管实验中低于10mmHgCO2 气腹对肝硬化兔肝脏血流影响较轻,但肝硬化患者行腹腔镜手术时仍需注意腹内压力和气腹持续时间。     

Electron-microscopic alterations of the peritoneum after both cold and heated carbon dioxide pneumoperitoneum

BACKGROUND: Carbon-dioxide (CO(2)) is used universally as an insufflation agent to create a laparoscopic pneumoperitoneum. In this study, we aimed to examine the electron and light microscopic alterations of the peritoneum after both cold-dry and heated-humidified CO(2) pneumoperitoneum. MATERIALS AND METHODS: Thirty male Sprague-Dawley rats were used in this study. The rats were separated into three groups each comprising 10 rats. Group-I: (Control group): Gas insufflation was not applied to these animals. Group-II: These animals received standard cold-dry (21 degrees C, 2% relative humidity) CO(2). Group-III: These animals received heated-humidified (40 degrees C, 98% relative humidity) CO(2). In groups II and III, peritoneal gas was emptied 2 h after pneumoperitoneum application. All rats were killed after 12 h. Peritoneal samples were examined both by scanning electron and light microscopy by two different pathologists who were not aware of the groups. RESULTS: According to light microscopic examination; in group II and III, cellular response (increased lymphocyte) was significantly higher than the control group (P < 0.01). Similarly, in group II cellular response was significantly higher than group III. (P < 0.01). There was no difference in increased capillarity among all groups. (P > 0.05). According to scanning electron microscopic examination, in group I, normal peritoneum was covered by a sheet of flat mesothelial cells densely covered with microvilli. No intercellulary clefts and no free basal lamina were detected. In group II, drastic alterations of the surface layer were seen. The mesothelial cells had extreme desquamation, and the basal membrane was clearly visible. In group III, the mesothelial cells had bulged up to the surface layer and retracted. Intercellulary clefts become visible, but the basal lamina was not seen. CONCLUSIONS: Electron and light microscopic examination revealed that heated-humidified CO(2) results in less peritoneal alteration than cold-dry CO(2.) Accordingly, we believe that heated-humidified CO(2) is more suitable for pneumoperitoneum application in laparoscopic surgery especially in selected cases.     

胃肠肿瘤CO2气腹切口种植的对比研究

目的　进一步探讨肿瘤细胞切口种植的原因。方法　选用人胃癌细胞株 Ｓ Ｇ Ｃ７９０１ 及人肠癌细胞株 Ｌｏ Ｖｏ，注入雄性 Ｓ Ｄ 大鼠腹腔内，气腹机产生腹腔持续性气腹，通过腹腔穿刺管（５ ｍ ｍ） 收集气体至培养瓶中，３７ ℃、体积分数为５ ％ Ｃ Ｏ２ 环境中培养７ 天后，鉴定是否有肿瘤细胞生长，另取腹腔冲洗液作阳性对照。结果　胃癌细胞持续性气腹测试组中，应用１０６ 个／ｍｌ 胃癌细胞，气腹压达３０ｍ ｍ Ｈｇ ，流量为５ Ｌ／ｍｉｎ ，持续６０ 分钟时，７５ ％ 培养瓶中有肿瘤细胞生长。而相同状态下，未见肠癌细胞生长。结论　相同状态下，胃癌细胞较肠癌细胞更易从切口逃逸，且与肿瘤细胞的数量、气腹压力及持续时间呈正相关。     

大鼠肺移植术后早期肺组织ICAM-1表达的变化

目的　研究大鼠肺移植术后早期肺组织细胞间粘附分子 1(ICAM 1)表达的变化。方法　用免疫组织化学 (SABC)法及设置内参照的半定量RT PCR方法 ,对移植肺获再灌注及通气 4h后肺组织的ICAM 1蛋白及其mRNA表达水平进行观察。实验组 (n =6 )肺经低钾右旋糖酐液 4℃保存12h后 ,行同种异体左肺原位移植。对照组 (n =6 )充分游离左侧肺动、静脉及支气管。免疫组织化学检测结果按阴性 (- )、可疑 (± )、弱阳性 ( )、阳性 ( )、强阳性 ( )进行记录。PCR产物电泳后扫描记录条带密度。结果　移植肺肺泡上皮及肺血管内皮细胞免疫着色显著较对照组深 (P<0 .0 1)。其ICAM 1mRNA与β actinmRNART PCR扩增产物相对密度值亦显著高于对照组 ,分别为 0 .873± 0 .0 44和 0 .44 2± 0 .0 37,P <0 .0 1。结论　肺移植术后早期肺组织ICAM 1表达上调 ,这种上调与ICAM 1mRNA增强有关。     

The impact of carbon dioxide pneumoperitoneum on liver regeneration after liver resection in a rat model

Background  

In recent years, laparoscopic hepatic resection is performed by an increasing number of surgeons. Despite many advantages of the laparoscopic procedure, it is unclear whether the pneumoperitoneum affects the postoperative liver regeneration after liver resection. The current study aimed to investigate the influence of a carbon dioxide (CO₂) pneumoperitoneum on liver regeneration in a rat model.     

Liver metastases are less established after gasless laparoscopy than after carbon dioxide pneumoperitoneum and laparotomy in a mouse model

BACKGROUND: Although the liver is the most frequent site of cancer recurrence after conventional open surgery for colorectal cancer, the effect of laparoscopic procedures with or without gas insufflation on the development of liver metastases is largely unknown. METHODS: Male BALB/C mice inoculated intraportally with colon 26 cells were randomized to undergo carbon dioxide pneumoperitoneum (n = 14), abdominal wall lifting (n = 14), or full laparotomy (n = 12), or to serve as control subjects without any procedures other than tumor inoculation (n = 13). RESULTS: The growth of liver metastases 14 days after surgery was enhanced after full laparotomy (p < 0.01) and pneumoperitoneum (p = 0.02), as compared with that in the control subjects, whereas there was no difference in the growth of liver metastases between abdominal wall lifting and the control condition (p = 0.99). CONCLUSIONS: These results suggest that the defense against liver metastasis is better preserved after the gasless procedure than after laparotomy and carbon dioxide pneumoperitoneum in the reported animal model.