Effects of Ozone Oxidative Preconditioning on Liver Regeneration after Partial Hepatectomy in Rats

ABSTRACT

Aim: Similar protective effect of ischemic and ozone oxidative preconditioning (OzoneOP) in hepatic ischemia–reperfusion (I/R) injury was demonstrated, providing evidences that both preconditioning settings shared similar biochemical mechanisms of protection. We investigated the effects of OzoneOP on liver regeneration after 70% partial hepatectomy (PHx) in rats. Methods: Rats were divided into three groups: PHx, I/R + PHx, and OzoneOP + I/R + PHx groups. Ozone (intraperitoneal, 1.2 mg/kg) was given to rats subjected to I/R and 70% hepatectomy daily five times before operation. At 24 hr and 48 hr after resection, samples were collected for the measurement of serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), tumor necrosis factor alpha (TNF-α), and interleukin-6 (IL-6). Moreover, liver regeneration rate, proliferating cell nuclear antigen (PCNA) labeling index, mitotic index, and histopathological examination were evaluated. Results: OzoneOP reduced liver injury determined by liver histology and serum transaminases. There was a rise in serum TNF-α and IL-6 levels in the I/R + PHx group whereas OzoneOP significantly decreased the rise in the level of TNF-α but not IL-6 on the 24 hr and 48 hr of reperfusion. Moreover, liver regeneration in OzoneOP + PHx group, as assessed by the regenerated liver weight, mitotic, and PCNA-labeling index, was significantly improved when compared to I/R + PHx group. Conclusion: These results suggest that OzoneOP ameliorates the hepatic injury associated with I/R and has a stimulatory effect on liver cell regeneration that may make it valuable as a hepatoprotective modality.     

生长抑素对鼠大部肝切除术后肝再生的影响

目的　探讨生长抑素对肝切除术后肝再生的影响。　方法　 6 0只SD大鼠随机分为对照组及生长抑素组 ,按Higgins方法行 70 %肝切除术 ,术后给药并分批于术后 6、2 4、48、72、96h处死 ,作如下比较 :①残肝肝重 ;②增殖细胞核抗原 (PCNA)标记指数 ;③图像定量分析测量PCNA阳性产物面积。　结果　与对照组比较 ,生长抑素组残肝肝重在术后 48、72、 96h显著降低 (P <0 .0 5 ) ,PCNA标记指数、PCNA阳性产物面积在术后 2 4、48、72h显著降低 (P <0 .0 5 ) ,且以 2 4、 48h最为明显 (P <0 .0 1)。　结论　生长抑素能抑制肝再生 ,且肝再生程度越强 ,抑制效应越明显     

Quantitative Gene Expression Analysis by cDNA Microarray During Liver Regeneration After Partial Hepatectomy in Rats

Purpose It is speculated that genetic regulation plays an important role during liver regeneration. We conducted this study to analyze quantitative gene expression during liver regeneration after partial hepatectomy.Methods Sixty male Wistar rats were randomly assigned into ten groups of six. One group of rats was killed preoperatively and the other nine groups were killed 2, 4, 6, 12, 24, 48, and 72h, and 5 and 7 days after 70% partial hepatectomy, respectively. The remnant livers were isolated for mRNA extraction. A mass survey of gene expression by cDNA microarray carrying 6144 polymerase chain reaction-amplified cDNA fragments was prepared by an arraying machine. The microarray images were scanned, digitized, and analyzed using a flatbed scanner.Results The variations in gene expression were classified into 72 different patterns including a pattern with a single peak, 2, 4, 6, 12, and 72h, and 5 and 7 days after partial hepatectomy. The other patterns included double peaks, enhancing trend, diminished trend, protruding curve, excavated curve, and various mixed types. Each category of gene expression pattern contained 40–218 different proto-oncogenes.Conclusion The quantitative gene expression profiles have important implications, warranting further investigation of the genetic mechanisms involved in the process of liver regeneration.     

Total Blood Exchange Suppresses the Early Stage of Liver Regeneration Following Partial Hepatectomy in Rats

Despite its obscure and short effect, plasma exchange (PE) remains a mainstay in the treatment of liver disease. However, the question still remains as to whether or not PE suppresses the regeneration of the liver because PE deprives patients of hepatotrophic factors. The effect of PE, which could be a total blood exchange (TBE) in a syngeneic setting, on liver regeneration following a 68% partial hepatectomy (PH) was investigated in rats. In Group 1, 20 ml of blood from normal rats was infused while native blood was removed at 6 and 12 h after PH. In Group 2, 20 ml of blood obtained from PH rats at the same time points was infused. The regeneration rate, labeling index of proliferating cell nuclear antigen (PCNA), and plasma hepatocyte growth factor (HGF) level were determined, and standard liver function tests performed at 24, 48, and 72 h. Although all liver function tests improved in Group 1 at 24 and 48 h, the regeneration rate was significantly impaired. Similarly, the PCNA labeling index was significantly lower in Group 1 than that in Group 2. The plasma HGF level was significantly reduced in Group 1 (6 h blood out versus blood in: 1.1 ± 0.5 vs. 0.1 ± 0.1 ng/ml, p < 0.05). TBE with normal blood following PH suppressed the early stage of liver regeneration, in part, because of the reduction of HGF even though the blood was purified.     

Convergence Process of Volumetric Liver Regeneration After Living-Donor Hepatectomy

Background  

We investigated the long-term profiles of liver regeneration after living-donor hepatectomy.     

Angiogenesis Inhibitor TNP-470 Can Suppress Hepatocellular Carcinoma Growth Without Retarding Liver Regeneration After Partial Hepatectomy

Purpose. We investigated the suppressive effect of the angiogenesis inhibitor TNP-470 on accelerated hepatocellular carcinoma (HCC) growth in the regenerating liver. Methods. After 70% partial hepatectomy (PH), AH-130 cells were injected into the portal vein of Donryu rats. A control group was given the vehicle only, and the treated group was given 10mg/kg TNP-470 subcutaneously every second day, from 24h after tumor implantation, seven times. On day 14, tumor growth was evaluated by the number of foci on the liver surface, liver weight, and the microvessel density of the tumor. Results. The number of foci was significantly less in the treated group (116.5 ± 103.1) than in the control group (319.3 ± 223.1) (P 0.05), as was microvessel density, which was 31.3 ± 14.0/mm² in the treated group and 61.2 ± 18.9/mm² in the control group (P 0.05). The liver tended to weigh less in the treated group (12.15 ± 1.28g) than in the control group (15.22 ± 5.35g). We also assessed whether TNP-470 retards liver regeneration. Seven days after 70% PH, the liver weight in the treated group was similar to that in the control group. Total bilirubin, serum glutamic oxaloacetic transaminase, and serum glutamic pyruvic transaminase were not higher in the treated group than in the control group. Conclusion. TNP-470 can suppress HCC growth without retarding liver regeneration after PH.     

大鼠肝移植后肝再生的实验研究

目的：探讨部分肝移植术后移植肝的再生问题。方法：建立大鼠部分肝移植模型，实验分为肝切除组（PLR组）、全肝移植组（OLT组）和部分肝移植组（POLT组）3组，分别于术后不同时间段取外周血检测总胆红素和谷丙转氨酶水平；取肝组织行组织学检查及流式细胞仪检测移植肝的增殖活性。结果：移植术后1w，肝功能酶学指标增高，后逐步降低；组织学检查术后可见单核细胞浸润，特别在门静脉周围汇管区，肝实质可见点状坏死。术后1个月可见胆管增殖；PLR组和POLT组还可见二倍体和多倍体的肝细胞，中央小静脉、肝窦和叶间静脉轻度扩张。PLR组和POLT组肝细胞增生活跃，3组分别于术后1d、2d、4d达到增殖高峰。结论：部分移植肝和肝切除后肝脏具有同样的增殖活性，但增殖高峰POLT组及OLT组均要晚于肝切除后的肝脏，但移植组增殖周期长。这可能是由于手术操作及肝脏缺血再灌注损伤所致。而持续时间长可能与受体免疫系统产生的细胞因子和激素的调控相关。     

Glycine Pretreatment Ameliorates Liver Injury After Partial Hepatectomy in the Rat

ABSTRACT

Background: Living donor liver transplantation subjects the donor to a major hepatectomy. Pharmacological or nutritive protection of the liver during the procedure is desirable to ensure that the remnant is able to maintain sufficient function. The aim of our study was to analyze the effects of pretreatments with α-tocopherol (vitamin E), the flavonoid silibinin and/or the amino acid L-glycine on the donor in a rat model. Methods: Male Wistar rats were pretreated with L-glycine (5%% in chow, 5 days), α-tocopherol (100 mg/kg body weight by gavage, 3 days) and/or silibinin (100 mg/kg body weight, i.p., 5 days). Thereafter, 90%% partial hepatectomy was performed without portal vein clamping. Results: Glycine pretreatment markedly decreased transaminase release (AST, 12 hr: glycine 1292 ± 192 U/L, control 2311 ± 556 U/L, p < .05; ALT, 12 hr: glycine 1013 ± 278 U/L, control 2038 ± 500 U/L, p < .05), serum ALP activity and serum bilirubin levels (p < .05). Prothrombin time was reduced, and histologically, liver injury was also decreased in the glycine group. Silibinin pretreatment was less advantageous and pretreatment with α-tocopherol at this very high dose showed some adverse effects. Combined, i.e., triple pretreatment was less advantageous than glycine alone. Liver resection induced HIF-1α accumulation and HIF-1α accumulation was also decreased by glycine pretreatment. Conclusion: The decrease of liver injury and improvement of liver function after pretreatment with glycine suggests that glycine pretreatment might be beneficial for living liver donors as well as for patients subjected to partial hepatectomy for other reasons.     

Assessment of Effect of Intraperitoneal Tacrolimus on Liver Regeneration in Major (70%) Hepatectomy Model After Experimental Pringle Maneuver in Rats

AimSmall-for-size grafts have become more important, especially in living donor liver transplants. The Pringle maneuver, used to reduce blood loss, and the immunosuppressive medications used to prevent graft rejection in liver transplants have different side effects on liver regeneration. We researched the effect of situations where tacrolimus and the Pringle maneuver were applied or not on liver regeneration in rats with partial hepatectomy.Material and MethodsThis study was completed with 35 Wistar Albino rats. The subjects were randomly divided into 5 groups: Group 1 had the abdomen opened and no other procedure was performed; Group 2 underwent a 70% hepatectomy; Group 3 underwent a 15-minute Pringle maneuver + 70% hepatectomy; Group 4 underwent a 70% hepatectomy + 5 days of 1 mg/kg/day intraperitoneal tacrolimus; and Group 5 underwent a 150 minute Pringle maneuver + 0% hepatectomy + 5 days of 1 mg/kg/day intraperitoneal tacrolimus. All rats were sacrificed on the seventh postoperative day, remaining liver tissue was weighed, and weight indices created. The remaining liver tissue was stained with phosphohistone H3 and the mitotic index calculated.ResultsThe groups that underwent the Pringle maneuver, 70% hepatectomy, and tacrolimus administration were compared with the control group in terms of mitotic index and weight index, but no statistically significant differences were identified.ConclusionSuppression of regeneration forms a risk after liver transplantation with small-volume grafts. As a result, research on the effect of tacrolimus combined with the Pringle maneuver is important, especially for transplantations using segmented liver grafts. In our study, we showed that the use of tacrolimus had no negative effect on liver regeneration.     

重组人生长激素在治疗肝硬变大鼠肝部分切除术后低白蛋白血症中的作用

目的探讨重组人生长激素(rhGH)在治疗肝硬变大鼠肝部分切除术后低白蛋白血症中的作用。方法30只大鼠按编号法随机分为正常对照组、肝硬变组(LC组)、肝硬变切肝组(LCH组)、PN组(于肝叶切除术后行PN治疗)及rhGH PN组(于肝叶切除术后行rhGH PN治疗)．每组6只。检测大鼠肝功能、血糖．用RT-PCR法检测肝组织ALB mRNA的表达．肝组织行Ki67免疫组化染色。结果与PN组比较．rhGH PN组血清ALP降低，ALB、血糖升高．肝组织ALBmRNA表达水平及肝组织Ki67指数均升高。结论重组人生长激素可以改善肝硬变大鼠肝部分切除后低白蛋白血症。     

Percentage of Future Liver Remnant Volume Before Portal Vein Embolization Influences the Degree of Liver Regeneration After Hepatectomy

Background

Clinical determinants of liver regeneration induced by portal vein embolization (PVE) and hepatectomy remain unclear. The aims of this study were to investigate how liver regeneration occurs after PVE followed by hepatectomy and to determine which factors strongly promote liver regeneration.

Methods

Thirty-six patients who underwent both preoperative PVE and major hepatectomy were enrolled in this study. Percentage of future liver remnant volume before PVE (%FLR-pre) was compared with the remnant liver volume after PVE (%FLR-post-PVE) and on postoperative day 7 after hepatic resection (%FLR-post-HR). Clinical indicators contributing to liver regeneration induced by both PVE and hepatectomy were examined by logistic regression analysis.

Results

PVE and hepatectomy caused a two-step regeneration. FLR-pre, FLR-post-PVE, and FLR-post-HR were 448, 579, and 761 cm³, respectively. The %FLR-pre was significantly associated with liver regeneration induced by both PVE and hepatectomy (r?=?0.63, p?<?0.0001). Multiple regression analysis showed that only %FLR-pre was independently correlated with posthepatectomy liver regeneration (p?=?0.027, odds ratio?=?13.8).

Conclusion

After PVE and the subsequent hepatectomy, liver regeneration was accomplished in a two-step manner. Liver regeneration was strongly influenced by the %FLR-pre.     

Regeneration of Liver Function Capacity After Partial Liver Resection is Impaired in Case of Postoperative Bile Leakage

Background

Postoperative bile leakage (PBL) is a major surgical complication after partial liver resection resulting in increased perioperative morbidity and mortality. Previous experimental studies have shown an adverse effect on functional liver regeneration, but there are still no data available concerning these effects in humans.

Materials and methods

A retrospective matched pair analysis was carried out comparing the postoperative course of 51 patients with PBL to 51 patients without PBL. The influence of PBL on actual liver function was determined by means of LiMAx, Indocyanine green plasma disappearance rate (ICG-PDR), and standard liver function tests.

Results

The analyzed groups were matched regarding demographic, preoperative, and operative data. Bilirubin, ICG-PDR and LiMAx showed delayed postoperative functional recovery after partial liver resection in the PBL group compared to the non-PBL group. LiMAx was the single parameter to assess differing liver regeneration continuously from the 3rd to 14th postoperative day between the groups. A subanalysis revealed an early recovery from impaired functional regeneration if the bile leakage was diagnosed until the 5th postoperative day (POD) or successfully treated until the 14th POD.

Conclusion

These are the first data evaluating the impact of PBL on functional regeneration after liver surgery. Liver regeneration after partial liver resection is impaired by PBL. However, a nearly normal recovery is possible if the bile leak is diagnosed early and immediately treated. Moreover, patients presenting with delayed functional recovery in the postoperative course are likely to have complications, e.g., bile leakage.

     

Recurrence of Hepatocellular Carcinoma in Noncirrhotic Liver After Hepatectomy

Background

Hepatocellular carcinoma in noncirrhotic liver (HCCNC) is rare. This tumor has a particular epidemiology and presentation, and it requires specific treatment, compared with HCC in cirrhotic liver. The aims of this study were to determine the survival and recurrence rates, prognostic factors, and optimum treatment of HCCNC and to propose a follow-up protocol for patients who have undergone surgery for HCCNC.

Methods

This study included 131 patients who underwent surgical treatment for HCCNC from January 1992 to December 2010. Survival and recurrence rates were evaluated, and the prognostic factors and characteristics of recurrence were analyzed. Pathologic characteristics of the tumors and the nontumoral liver were examined.

Results

The mean survival time was 67.9 months. The 5- and 10-year overall survival rates were 72.9 and 36.7 %, respectively. In all, 54 patients (41.2 %) developed recurrence at a median interval of 30.96 months. Of these recurrences, 31.5 % occurred during the first year, and 24.1 % occurred more than 5 years after surgery. Macro- or microvascular invasion and tumor size >5 cm were significantly associated with a poor survival rate. The predictive factors for recurrence were multiple tumors, tumor diameter >5 cm, and satellite nodules. Patients who underwent surgical treatment for recurrence had a significantly longer survival time than those who did not (p < 0.0292).

Conclusions

Recurrence is the most common cause of death after hepatectomy for HCC, and patients should undergo careful, long-term follow-up. Early detection and treatment of recurrence with curative intent should improve the prognosis of these patients.     

S-腺苷蛋氨酸在肝硬变大鼠行部分肝叶切除后对肝功能的保护作用观察

目的　探讨S 腺苷蛋氨酸在肝硬变大鼠行部分肝叶切除后对肝细胞的再生及肝功能的影响。方法　用40 %四氯化碳诱导Wistar大鼠肝硬变模型 ,而后将成模大鼠随机分为肝硬变组 (n =2 0 )、治疗 1组〔S 腺苷蛋氨酸 10mg/ (kg·d) ,n =16〕及治疗 2组〔S 腺苷蛋氨酸 2 0mg/ (kg·d) ,n =16〕 ,另设正常对照组 (n =16)。 4组大鼠均行 3 0 %左右肝叶切除。自手术当天起 ,治疗 1、2组分别肌注S 腺苷蛋氨酸 10mg/ (kg·d)和 2 0mg/ (kg·d) ,对照组肌注生理盐水 5ml/d ,共 15d。 4组大鼠分别于术后 15d和 3 0d处死一半 ,取血检测Alb、ALT、TB、TBA及TNF α ,同时在光镜及电镜下观察肝组织的病理改变及超微结构变化。结果　肝硬变组术后 15d和 3 0d的TB、TBA、ALT及TNF α水平明显高于正常对照组 (P<0 .0 1) ,Alb水平明显低于正常对照组 (P<0 .0 1) ;治疗 1、2组术后 15d的上述 4项指标高于正常对照组 (P<0 .0 5 ) ,但明显低于肝硬变组 (P<0 .0 1) ,Alb水平低于正常对照组 (P<0 .0 5 ) ,却明显高于肝硬变组 (P<0 .0 1) ;治疗 1、2组术后 3 0d上述各指标均接近正常对照组水平。TNF α和TB、TBA、ALT呈显著正相关 (r值分别为 0 .99、0 .97、0 .93 ,P<0 .0 1) ,与Alb呈显著负相关 (r值为 -0 .88,P<0 .0 1)。治疗组光镜     

Platelets Promote Liver Regeneration in Early Period after Hepatectomy in Mice

Background Platelets contain several growth factors, including platelet-derived growth factor and hepatocyte growth factor. Materials and methods We examined the effects of platelet increment on liver regeneration after 70% hepatectomy. Hepatectomies were carried out in male BALB/c mice, and subsequently divided into three groups: (i) untreated mice, (ii) thrombocytotic mice induced with thrombopoietin, and (iii) thrombocytopenic mice induced with anti-platelet antibody. Growth kinetics in the liver were analyzed as a function of the liver/body weight ratio, the mitotic index, the proliferating cell nuclear antigen labeling index and Ki-67 labeling index. Activation of signal transduction pathways relating to cell proliferation were examined, including the STAT3, Akt, and ERK1/2 pathways. Platelet accumulation in the residual liver was quantified by immunohistochemistry and transmission electron microscopy. Results In thrombocytotic and thrombocytopenic mice, liver/body weight ratios and Ki-67 labeling indices were significantly increased and significantly decreased, respectively, compared with untreated mice 48 hours post-hepatectomy. The Akt pathway was strongly activated, and platelet accumulation was significantly increased in thrombocytotic group 5 minutes post-hepatectomy compared with normal and thrombocytopenic groups. After hepatectomy platelets accumulated in the sinusoids of liver and promoted hepatocyte proliferation in early period after hepatectomy. Conclusion By increasing or decreasing the platelet, marked changes in liver regeneration can occur, due to differences in cellular signaling and mitosis.