Kupffer cell-mediated inhibition of liver regeneration after combined hepatectomy and pancreatectomy

Recently, simultaneous hepatectomy and pancreatoduodenectomy has been performed for the treatment of some biliary tract cancers in Japan. Postoperative hepatic failure is a common and potentially fatal complication. The aim of this study was to examine the reduction in the rate of liver regeneration after 70% hepatectomy (Hx) alone or in combination with 70% pancreatectomy (HPx). Male Sprague-Dawley rats underwent hepatectomy or simultaneous hepatectomy and pancreatectomy. The ratio of liver weight to body weight, the labeling index of hepatocytes in vivo, and DNA synthesis of the hepatocytes and/or Kupffer cells in primary culture were analyzed. The ratio of liver weight to body weight and the labeling index in HPx rat were found to be significantly lower than those values in Hx rats. There were no significant differences in plasma alanine aminotransferase levels between the two groups. The inhibitory effect on DNA synthesis was observed with coculture of hepatocytes and Kupffer cells when the portal plasma obtained 1 hour after operation was added. We further observed that the conditioned medium of Kupffer cells stimulated by the addition of the portal plasma that was obtained 1 hour after HPx inhibited DNA synthesis of hepatocytes. This effect was abolished after incubation at 56° C for 30 minutes. These results strongly suggest the existence of a growth inhibitory factor in portal plasma after HPx. This heat-labile growth inhibitory factor was released from Kupffer cells and would appear to act on hepatocytes in a paracrine manner. Supported by the Kanae Foundation for Life and Sociomedical Science, Japan. Presented at the Thirty-Eighth Annual Meeting of The Society-for Surgery of the Alimentary Tract, Washington, D.C., May 11–14, 1997.     

Effect of LAK cells on liver regeneration after partial hepatectomy

Lymphokine activated killer (LAK) cells can destroy not only tumor cells but also syngeneic regenerating liver cells. This study was started to determine the effect of passive transfer of LAK cells on liver regeneration after partial hepatectomy. C3H mice were received 70% hepatectomy and LAK cells were injected intravenously at a dose of 5 X 10(7) cells/body. After 36 hours, 3H-thymidine uptake into the residual liver was measured. LAK cells transferred group showed 31% suppression compared with control group. In vitro, 24 hours addition of LAK cells to the primary culture of regenerating liver cells caused 97% suppression of 3H-thymidine uptake at effector to target ratio, 50/1. Then we examined the effects of IL-2 administration on liver regeneration. Though IL-2 showed no effect on cultured liver cells, intraperitoneal administration of IL-2 after hepatectomy at a dose of 1 X 10(4)u/body 5 times every 8 hours brought 38% suppression of 3H-thymidine uptake of the residual liver. Cyclosporine A, which can suppress the IL-2 production of lymphocytes, promoted liver regeneration 45% over the control at a dose of 10 mg/kg. These results suggest that LAK cells could regulate liver regeneration.     

肝再生增强因子促进肝再生的细胞信号途径研究

目的探讨肝再生增强因子（ALR）促进肝部分切除（PH）后再生的机制。方法采用肝再生率评价ALR对PH后肝再生的作用；MTT检测ALR协同肝再生大鼠血清对肝细胞增殖和ALR对TGF-β1抑制肝细胞增殖的作用；免疫组化观察再生肝中ALR的表达；免疫细胞化学和Western blot检测经ALR刺激后肝细胞中ALR表达变化。结果ALR明显促进PH后的肝再生，可协同肝再生大鼠血清促进肝细胞增殖；ALR能拮抗TGF-β1对肝细胞增殖的抑制；ALR主要表达于正常肝细胞浆内；PH后2～3d肝细胞ALR减少甚至消失，第6天随着肝再生的完成，ALR含量又恢复正常；ALR刺激后肝细胞内源性ALR表达增高。结论ALR能以自主分泌方式从肝细胞中释放，与间质细胞产生的肝再生调控因子相互作用，如抑制TGF-β1生物活性，间接促进PH后的肝再生过程。     

Functional assessment of liver regeneration after major hepatectomy

BackgroundLiver regeneration is crucial to restore the functional liver mass after liver resection. The aim of this study was to evaluate the early postoperative changes in remnant liver function, volume and liver stiffness after major liver resection and their correlation with postoperative outcomes.MethodsPatients undergoing major liver resection (≥3 segments) between February and November 2018 underwent both functional assessment using technetium-99m mebrofenin hepatobiliary scintigraphy (HBS) and CT-volumetry of the (future) remnant liver on preoperative day 1, the 5^th postoperative day, and 4–6 weeks after resection. At the same time points, patients underwent transient elastography (TE) for the assessment of liver stiffness. Severe postoperative complications (Clavien-Dindo ≥ 3A) and mortality were correlated with the functional and volumetric increases of the remnant liver. Liver failure was graded according to the International Study Group of Liver Surgery (ISGLS) criteria.ResultsA total of 18 patients were included of whom 10 (56%) had severe complications and one patient (5%) developed liver failure. Function and volume of the remnant liver had increased by the 5^th postoperative day from 6.9 (5.4–10.9) to 9.6 (6.7–13.8) %/min/m², P=0.004 and from 795.5 (538.3–1,037.5) to 1,080.0 (854.0–1,283.3) mL, P<0.001, respectively. After 4–6 weeks, remnant liver volume had further increased [from 1,080.0 (854.0–1,283.3) to 1,222.0 (1,016.0–1,380.5) mL, P=0.035], however, liver function did not show any significant, further increase [from 9.6 (6.7–13.8) to 10.9 (8.8–13.6) %/min/m², P=0.177]. Liver elasticity of the future remnant liver (FRL) increased [from 10.8 (5.7–18.7) to 17.5 (12.4–22.6) kPa, P=0.018] and gradually recovered after 4–6 weeks to a median of 10.9 (5.7–18.8) kPa (T3 vs. T4, P=0.079). Patients who had severe postoperative complications did not show a significant increase in liver function on the 5^th postoperative day (P=0.203), despite increase of volume (P<0.01).ConclusionsFunctional regeneration of the remnant liver predominantly occurs during the first 5 days after resection. In case of severe complications, functional regeneration is delayed, in contrast to volume increase.     

索拉非尼对肝癌术后早期肝再生的影响

目的研究索拉非尼对肝癌患者术后早期肝再生、肝功能及凝血功能的影响,以及癌旁肝组织的血管内皮生长因子受体2（VEGFR-2）、血管内皮生长因子受体3（VEGFR-3）、血小板衍生的生长因子受体β（PDGFR-β）和原癌基因丝苏氨酸蛋白激酶1（C-Raf-1）的表达水平与肝再生的关系。方法 30例经病理确诊为肝细胞癌的患者,手术切除后接受不同剂量索拉非尼治疗。应用CT测量术后肝脏体积变化,比较不同剂量的索拉非尼对术后1、3个月时肝再生率的影响。分析不同剂量下,服药前及服药后1个月、3个月时患者肝功能及凝血功能的变化。通过免疫组化检测癌旁肝组织中VEGFR-2、VEGFR-3、PDGFR-β和C-Raf-1的表达,分析表达水平与术后1、3个月时肝再生率的相关性。结果索拉非尼服药剂量400 mg/d组的患者术后1个月、3个月时肝再生率平均增加量高于800 mg/d组的患者（P〈0.05）。两组患者服药后3个月时血清AST、ALT明显比服药前升高,血清凝血酶原时间（PT）明显延长（P〈0.05）。术后1、3个月时肝再生率与VEGFR-2、PDGFR-β表达呈负相关（P〈0.05）,VEGFR-2表达对肝再生率的影响最大。结论索拉非尼对肝癌术后早期肝再生有抑制作用,剂量越大抑制效果越明显。VEGFR-2是索拉非尼抑制早期肝再生的主要影响因素。     

Plasminogen mediates liver regeneration and angiogenesis after experimental partial hepatectomy

BACKGROUND: Plasmin system components are upregulated after partial hepatectomy, but their contribution to surgery-induced hepatic angiogenesis and regeneration is unclear. Liver regeneration and angiogenesis after partial hepatectomy were examined in mice lacking plasminogen or urokinase plasminogen activator (uPA). METHODS: Mice with a single-gene deletion of plasminogen or uPA were subjected to 70 per cent partial hepatectomy. Liver regeneration was measured as relative liver weight and cell proliferation index. Angiogenesis was quantified by determining hepatic microvessel density after staining for sinusoidal endothelial cells. RESULTS: The liver remnant weight was significantly reduced in mice lacking plasminogen or uPA compared with that in wild-type mice on days 2 and 7 after partial hepatectomy. This correlated with impaired cell proliferation. In wild-type mice, regeneration was accompanied by a significant increase in microvessel density after hepatectomy; this increase was impaired in plasminogen-deficient mice. CONCLUSION: Plasminogen and uPA are essential for optimal liver regeneration. In addition, plasminogen appears to be a major determinant in regeneration-associated hepatic angiogenesis.     

Sildenafil accelerates liver regeneration after partial hepatectomy in rats

Objective

The regeneration process causes the liver to achieve an adequate volume and function after major hepatectomy or living donor liver transplantation. Sildenafil, a selective phosphodiesterase-5 inhibitor used for erectile dysfunction, impacts the liver by enhancing the effects of nitric oxide. The aim of this study was to investigate the influence of sildenafil on liver regeneration in rats after partial hepatectomy.

Methods

Sixty young female Wistar Albino rats were randomly divided into three equal groups before 70% hepatectomy. Thereafter, we administered intraperitoneal saline to the control group (G1); 10 μg/kg sildenafil to the low-dose group (G2) and 100 μg/kg to the high-dose sildenafil group (G3). Half of the rats per group were sacrificed on the first and the other half on the fifth postoperative day after partial hepatectomy. Regeneration was assessed using three methods: (1) the formula described by Kwon et al formula, (2) the average number of mitotic figures in 10 microscopic fields, and (3) the average of Ki-67-positive nuclei in 1000 cells using immunohistochemistry.

Results

Although, the hepatic regeneration and mitosis rates were similar in all three groups, Ki-67 levels were significantly higher in both G2 and G3 than the control group on the first postoperative day. Hepatic regeneration was significantly greater in G2 and G3 than the control group as was the mitosis rate in the G2 group versus the two groups. By the 5th postoperative day Ki-67 levels were similar in the three groups.

Conclusion

Sildenafil treatment accelerated hepatic regeneration after partial hepatectomy in rats.     

两种肝再生模型肝切除术后肝再生指数和肝再生度的实验研究

【摘要】目的　探讨大鼠肝细胞增殖介导肝再生和卵圆细胞增殖介导肝再生两种不同大鼠肝再生模型肝切除术后肝再生指数和肝再生度变化的情况。方法　SD 大鼠随机分为2组: ①肝细胞增殖介导肝再生模型组(PH)②卵圆细胞增殖介导肝再生模型组(2AAF/PH) ,计算两组模型肝切除术后4、8、12、16、20天残肝肝再生度及肝再生指数。结果　两组肝再生度、肝再生指数均在第4天明显升高,12天左右达到高峰,超过再生总量的2/3以上,至20天基本达到原肝重。在4、8、12天同一时间点内,PH组肝再生度比2AAF/PH组高（P<0.05）,而到16天时,2AAF/PH组肝再生度则比PH组高（P<0.05）。在4、8天同一时间点内,PH组肝再生指数比2AAF/PH组高（P<0.05）。在12、16、20天同一时间点内,两组肝再生指数无显著性差异(P>0.05)。结论　肝再生度与肝再生指数是评价肝再生质量变化较直观和准确的指标,肝再生度在反映肝再生规律方面要比肝再生指数更准确。     

VEGF is important for early liver regeneration after partial hepatectomy

BACKGROUND: The aim of the study was to determine the role of Vascular Endothelial Growth Factor (VEGF) on the microvasculature and on angiogenetic gene expression after partial hepatectomy (PH) in the rat model. METHODS: To determine the effect of exogenous and endogenous VEGF after PH, rats were subjected to 70% PH and treated either with VEGF, anti-VEGF or NaCl. Postoperatively (3-168 h), vessel density (VD), vessel diameter (VDi), and intersinusoidal space, liver body weight ratio (LBR), hepatic proliferation and biochemical markers were assessed. To further elucidate the underlying molecular mechanisms hepatic gene expression was determined by customized cDNA arrays and quantitative RT-PCR. RESULTS: In the VEGF group, VD, VDi, and LBR were significantly increased compared with anti-VEGF or controls. Blockage of endogenous VEGF led to a marked increase of biochemical markers. Anti-VEGF almost completely suppressed and VEGF markedly enhanced hepatic proliferation in the first 24 h after surgery. This was associated with a modulation of cell cycle control genes (PC4, Gadd45a, Tis21/BTG2), v-jun, and CD14 by VEGF. CONCLUSIONS: VEGF plays an important role in liver regeneration and this may be due in part through its effects on neovascularization. Whether it may, when given therapeutically, represent a strategy to optimize liver regeneration in problematic patients needs to be clarified.     

Effect of splenectomy on liver regeneration and polyamine metabolism after partial hepatectomy

The effect of splenectomy on hepatic ornithine decarboxylase (ODC) induction, DNA synthesis, and mitosis of hepatocytes was studied in rat liver after partial hepatectomy. ODC activity markedly increased in the early stages of liver regeneration, and the increase in the activity was significantly enhanced in splenectomized rats. Splenectomy specifically induced ODC since tyrosine aminotransferase and general protein synthesis were not affected. Splenectomy also enhanced increase in hepatic polyamines, DNA synthesis, and mitosis in regenerating liver. The results suggest that splenectomy affects liver regeneration after partial hepatectomy by enhancing induction of ODC activity, which is an important biochemical event in the early stage of liver regeneration.     

肝脏类器官构建及其对小鼠部分肝切除术后肝再生的作用#br#

目的 观察肝脏类器官对小鼠部分肝切除术后肝再生的作用。方法 体外构建肝脏类器官,通过形态学、PCR和免疫荧光对类器官进行初步鉴定。C57BL/6小鼠随机等分为对照组和治疗组,每组18只。对照组进行肝左叶、中叶切除术+肝包膜注射200 μL PBS;治疗组进行肝左叶、中叶切除术+肝包膜移植200 μL类器官悬液。建模成功后分别于术后第1、4、7天收集标本。通过血清生化检测、免疫组化和Western blotting评估肝脏类器官对小鼠肝部分切除术后肝再生的作用。结果 类器官体外培养3 d,从直径20 μm的囊性结构扩增到约125 μm的细胞团（P＜0.05）,直径扩增近6 倍。肝脏干细胞标志基因EPCAM、SOX9和CK19明显上调（P＜0.05）,传代前后基因保持稳定。免疫荧光显示CK8、Desmin、AFP和PCNA呈阳性。HE显示术后第4天,治疗组肝细胞形态大小基本恢复正常,形态较清晰,无炎症细胞浸润,无脂肪或气球样变。对照组小鼠肝细胞核仁染色加深,仍有炎性细胞浸润,部分区域存在肝细胞坏死区。免疫组化Ki67和Western blotting对增殖水平进行检测,结果显示治疗组增殖能力是对照组的3 倍。肝功能检测发现治疗组在术后第4天ALT、AST、TBIL和DBIL明显下降,ALB合成增加（P＜0.05）。结论 具有肝脏干细胞属性和增殖能力的肝脏类器官,能通过保护肝细胞、改善部分肝切除术后小鼠肝功能,发挥促进肝再生作用。     

Increased intraabdominal pressure impairs liver regeneration after partial hepatectomy in rats

BACKGROUND: There are many experimental studies showing that increased intraabdominal pressure (IAP) reduces liver blood flow, leading to ischemia and portal venous congestion. But, there is no study evaluating the effect of increased IAP on liver regeneration. It is well known that acute liver ischemia and portal venous congestion impair liver regeneration. We, therefore, aimed to determine the effect of increased IAP on liver regeneration in this study. METHODS: Sprague-Dawley rats underwent partial hepatectomy with or without IAP of 12-14 mm Hg for 24 h or sham operation. Rats were randomly divided into six groups: two sham-operated groups, two hepatectomy groups, and two hepatectomy with increased IAP groups. Mitotic index, proliferating cell nuclear antigen (PCNA)-labeling index, and liver regeneration rate as liver regeneration parameters were studied on day 1 or on day 4 after operation. Additionally, serum aspartate transaminase (AST) level and histopathological changes in intestinal mucosa were studied. RESULTS: Hepatectomy with/without increased IAP groups had significantly higher serum AST levels than the sham-operated group on day 1. Serum AST level was found to be significantly higher in the hepatectomy with increased IAP group than in the other groups on day 4. Intestinal mucosal injury was found in the hepatectomy with increased IAP groups on days 1 and 4. Mitotic index and PCNA-labeling index were markedly higher in all hepatectomy with/without increased IAP groups than in the sham-operated groups. However, together with liver regeneration rate, both indices were significantly less in the hepatectomy with increased IAP groups than in the hepatectomy groups both on day 1 and on day 4. CONCLUSION: Maintenance of IAP between 12 and 14 mm Hg for 24 h impaired liver regeneration after partial hepatectomy in rats.     

Impact of hepatic vein deprivation on liver regeneration and function after major hepatectomy

Background and aims In extended liver resections, the preservation of vascular and biliary structures of the entire remnant liver is of paramount importance. The impact of venous outflow impairment and its consequences for liver regeneration and function are still a matter of debate. Materials and methods Rats (n = 75) were subjected to a 90% partial hepatectomy (PH), to a 70% liver resection with narrowing of the hepatic outflow of an additional 20% parenchyma (70%+ PH) or to an anatomic 70% PH. Postoperatively hepatocyte proliferation (Ki-67), liver function and survival were assessed. Gene expression analysis for markers of regeneration was determined by in-house complementary (DNA) arrays and quantitative real-time polymerase chain reaction (RT-PCR). Results Ninety percent PH led to a greater regenerative response as shown Ki-67 compared to animals with a 70%+PH (p < 0.05). However, liver function was equally impaired in both groups. Rats with 70% PH showed a greater proliferation index with less hepatic injury and better liver function. While mortality was 0% in the group of 70% PH, rats with 90% PH and 70+PH had a reduced survival of 75% (p < 0.05) Conclusion Venous outflow obstruction leads to an impairment of liver regeneration and liver function. In cases with critically small liver remnants, restoration of an adequate venous outflow may be mandatory.     

肝部分切除术后肝脏再生调控的研究进展

肝部分切除术(PH)后肝脏能迅速表现出巨大的再生能力.关于肝再生能力的研究一直是肝脏外科领域的热点.现已发现肝脏再生是一个涉及多种效应细胞增殖反应的高度协调的过程,其调控机制非常复杂.笔者就肝部分切除术后其再生调控的研究进展作一简要综述.