To date, however, a few investigations have been done on the potential use of MSCs to improve the outcome of small for size liver transplantation (SFSLT). The therapeutic mechanisms of MSCs are also not clearly elucidated. Whether MSCs contribute to liverregeneration by trans-differentiation into liver cells or by paracrine effects of their trophic factors has been ongoing discussions . Both Zhong Z and our previous study have already confirmed that defective liverregeneration in small grafts was associated with an inhibition of the c-Jun N- terminal kinase (JNK/c-Jun) and cyclin D1 (CyD1) pathways and compromised energy production [4, 5]. We also found that MSCs-conditioned medium could reduce liver injury and enhance regeneration in 50% reduced-size rat liver transplantation . Therefore, in this study we sought to determine whether and how MSCs therapy promoted liverregeneration and subsequently prolonged the rat survival in 30% SFSLT.
To characterize the function of LncPHx2, we developed two ASOs that efficiently target LncPHx2 for degradation by RNase H-mediated mechanism [31, 32](S2 Fig). We treated mice subcutaneously with LncPHx2_ASOs, and then performed PHx surgery as shown in Fig 3A. LncPHx2_ASO1 treatment completely blocked the upregulation of LncPHx2 during the liverregeneration process (Fig 3B). Interestingly, mice treated with LncPHx2_ASO1 had more rapid liver weight recovery compared to PBS-treated mice (Fig 3C). The effect on liver weight was significant at 48 hours after PHx, and livers in LncPHx2_ASO1-treated mice remained heavier 15 days after PHx (Fig 3C). The body weights of LncPHx2_ASO1-treated mice were not significantly different from PBS_treated mice (data not shown). The same phenotype was observed with two different ASOs targeting LncPHx2, but not with scrambled control ASO (S3 Fig). More proliferating cells were observed at 36 and 60 hours after PHx in LncPHx2_ASO1-- treated mice than in controls, as demonstrated by higher percentages of Ki67 staining and BrdU labelling (Fig 3D). No measurable differences in cell proliferation between LncPHx2_A- SO1-treated and PBS-treated mice were observed at later time points (Fig 3D). qPCR analyses showed elevated expression of cell-cycle marker genes in LncPHx2_ASO1-treated mice includ- ing genes encoding Cyclin E1 (Ccne1), Cyclin B1 (Ccnb1), and Aurora Kinase B (Aurkb) at 36 hours after PHx, and Mini-chromosome maintenance complex component 3 (Mcm3) at both 36 and 48 hours after PHx (Fig 3E). Ccnyl1 mRNA levels did not change upon LncPHx2-deple- tion in either regenerating livers or in livers subjected to sham surgery (data not shown). In addition, we observed a faster recovery of liver aspartate transaminase (AST) levels in plasma
Figure 1. Evaluation of liverregeneration in IL-1ra ko mice and in WT mice after 70% hepatectomy. The evaluation of hepatocyte proliferation on native liver tissue was performed by analyzing BrdU incorporation by immunohistochemistry and by analyzing PCNA expression by western blot analysis at 24 h, 48 h, 72 h, 5 days and 7 days after 70% hepatectomy in WT mice and IL-1ra KO mice. (A) BrdU incorporation was significantly higher in WT mice compared to IL-1ra KO mice at 24 h and 48 h after 70% hepatectomy. The proliferation index of BrdU-stained tissue was determined at 200x magnification and labelled nuclei were counted in 5 randomly chosen fields, which approximate 1000 cells per section. Data were expressed as the percentage of BrdU-stained hepatocytes per total number of hepatocytes. Positive hepatocytes for BrdU incorporation are stained in brown. (Original magnification x200). (B) For WT mice, PCNA expression peaked at 48 h, in contrast for IL-1ra KO mice, the PCNA expression peaked at 72 h. The results are shown as a ratio of PCNA to actin expression. The quantification of signals of PCNA and actin are performed by densitometry. Scale bars = 50 mm. * = Statistical significance p,0.05. D = day, WT = wild type, KO = knock-out, PCNA = proliferating cell nuclear antigen.
Rapidly proliferating tissue may require enhanced DNA repair capacity in order to avoid fixation of promutagenic DNA lesions to mutations. Partial hepatectomy (PH) triggers cell proliferation during liverregeneration (LR). How- ever, little is known on how DNA repair genes change and how they are regulated at the transcriptional level during LR. In the present study, the Rat Genome 230 2.0 array was used to detect the expression profiles of DNA repair genes during LR, and differential expression of selected genes was confirmed by real-time RT-PCR. 69 DNA repair genes were found to be associated with LR, more than half of which distributed in a cluster characterized by a grad- ual increase at 24-72h and then returning to normal. The expression of base excision repair- and transcrip- tion-coupled repair-related genes was enhanced in the early and intermediate phases of LR, whereas the expression of genes related to HR, NHEJ and DNA cross-link repair, as well as DNA polymerases and related accessory factors, and editing or processing nucleases, were mainly enhanced in the intermediate phase. The expression changes of genes in DNA damage response were complicated throughout the whole LR. Our data also suggest that the expres- sion of most DNA repair genes may be regulated by the cell cycle during LR.
Liverregeneration is a well-orchestrated phenomenon essential for both acute restoration of liver volume after resection and maintenance of its volume during chronic liver injury. A regenerating liver, after partial hepatectomy (PH), requires the formation of a complex net- work of liver sinusoids through which the blood flows. Thus, the angiogenic phase is a funda- mental process of LR, occurring predominantly in the later phase . Among the seven members of the vascular endothelial growth factor (VEGF) family (VEGF-A, VEGF-B, VEGF-C, VEGF-D, VEGF-E, VEGF-F, and PlGF), VEGF-A lies at the forefront of blood ves- sels formation[2, 3]. VEGF-A is known as the most potent and specific growth factor for both angiogenesis and vasculogenesis[4, 5]. Importantly, VEGF-A holds a regulatory role in differ- entiation and growth of liver sinusoidal endothelial cells (LSECs) . VEGF-A has been con- sidered as a central angiogenic player of LR.
nuclear antigen-labeling index, by immunohistochemical staining, was similar for both LED-exposed groups (P > 0.05) and higher than for the control group (P < 0.05). The cell proliferation index obtained with LED and laser were similar (P > 0.05). In conclusion, the present results suggest that LED irradiation promotes biological stimulatory effects during the early stage of liverregeneration and that LED is as effective as laser light, independent of the coherence, divergence and cromaticity.
Lipid metabolism was another pathway that was significantly impacted by A20. It is well established that mobilization of peripheral fat, hepatic FA uptake, and rapid accumulation of intracellular triglycerides in hepatocytes are important for optimal liverregeneration [45,46]. Cytokine signals after liver injury or LR promote the release of FA into the circulation and their uptake by hepatocytes . As a result, lipid droplets form in hepatocytes during the early stages of liverregeneration, providing energy to fuel the proliferative response. Even though this transient steatosis is part of the physiologic proliferative response, when excessive or uncontrolled, it can impair liverregeneration [48,49]. Several signals and molecules contribute to lipid accumulation during the regenerative process including Leptin signaling , hepatic Glucocorticoid-Receptor signaling, and Caveolin-1[51,52]. Our results show no evidence for A20 affecting the expression of Caveolin-1 or the Glucocorticoid-receptor complex, but obviates that overexpression of A20 significantly inhibits the up-regulation of the Leptin-receptor following 78% LR (31-339, Suppl. Table 2). This points to an intriguing regulation of this system by A20 and supports the hypothesis of Newberry et al and Leclercq et al, that Leptin signaling per se is not directly involved in liverregeneration, but rather required to promote the inflammatory and priming response, following PH [50,53]. Accordingly, when the priming response and the energy stores are already amplified by A20, we surmise that upregulation of the Leptin receptor is no longer required.
Figure 2. Augmenter of liverregeneration (ALR) regulated the proliferation of U266 multiple myeloma (MM) cells. A, U266 MM cells were exposed to 20 or 40 mg/mL recombinant ALR protein for 24–72 h and cell viability was examined by the MTT assay. ALR treatment signiﬁcantly promoted the proliferation of U266 MM cells. *Po0.05 vs untreated control cells; n=6 (one-way ANOVA with the Tukey test). B, qRT-PCR analysis of the ALR mRNA level in U266 cells transfected with control or ALR shRNA; n=3. C, MTT assay was done to measure the viability of U266 cells transfected with control or ALR shRNA after culturing for 72 h; n=6. D, Cell proliferation was determined by BrdU incorpora- tion assay after culturing for 72 h. ALR silencing signiﬁcantly suppressed the proliferation of U266 MM cells. *Po0.05 vs the control shRNA group; n=6 (Student t-test).
Tacrolimus) in the first 24 hours and on the seventh post- hepatectomy day. When the values were compared within each group there was an increase in both, more pronounced in the study group. A study similar to that, also using the Kwon formula to calculate the rate of liverregeneration in rats, revealed that the remaining liver had higher volume in both groups (control/experiment) both within 24 and seven days, but without statistical significance. The experiment group received glutamine supplementation at a dose 50ìg/day orally 7 .
day). The control group was not treated before partial hepatectomy and recovered under normal ambient conditions after the procedure. Groups postHBO and postHB were treated after partial hepatectomy with HBO and HB, respectively, three times (45 min/day). The preHBO group presented a significant increase in the initiation of the regenera- tion process of the liver 54 h postoperatively. The liver/body weight ratio was 0.0618 ± 0.0084 in the preHBO compared to 0.0517 ± 0016 g/g in the control animals (P = 0.016). In addition, the preHBO group showed significant better liver function (evaluated by the lowest serum ALT and AST activities, P = 0.002 and P = 0.008, respectively) and showed a significant decrease in serum albumin levels compared to control (P < 0.001). Liver lipid peroxide concentration was lowest in the preHBO group (P < 0.001 vs control and postHBO group) and light microscopy revealed that the composition of liver lobules in the preHBO group was the closest to normal histological features. These results suggest that HBO pretreatment was beneficial for rat liverregeneration after partial hepatectomy.
When using the Ki-67 index (percentage of labeled cells / total analyzed cells), there was no significant difference between the two 24 hour subgroups (CG 24h X SG 24h ). In the survey there was a statistically significant difference when compared the seven-day subgroups (CG 7d X SG 7d – p = 0.04). At the end of seven days of liverregeneration, the study group had a higher percentage of labeled hepatocytes when compared with the control group, demonstrating a greater number of hepatocytes in the group using sirolimus. This finding could suggest a stimulus on liverregeneration. Nevertheless, the peak of production of DNA followed by mitosis occurs early, and even if subsequent peaks take place, these are less
The liver’s ability to regenerate in mammals is a very well studied response. When toxic injury, exposure to vi- ruses, trauma or surgical resection results in loss of hepatic tissue, the remnant liver lobes will compensate for lost tis- sue and recover the initial liver mass within two weeks (Fausto et al., 2006). In the case of partial hepatectomy (PH), it results in hypertrophy of the remnant liver rather than restoration of the resected lobes as a consequence of cell proliferation, with the goal of replacing lost functional mass, which is called liverregeneration (LR) (Higgins and Anderson, 1931). The degree of hyperplasia is precisely controlled by the metabolic needs of the organism, prolifer- ating under conditions of functional deficiency, and under- going apoptosis under functional excess, such that the process stops once an appropriate liver to body weight ratio is achieved. In mouse and rat, the ratio is about 4.5%; in hu- mans this number is approximately 2.5% (Fausto, 2000; Riehle et al., 2011). PH triggers the actions of signaling pathways, growth factors and cytokines, cell cycle associ- ated proteins, and extracellular matrix, etc., leading to cell proliferation and structure function reorganization (Pahla- van et al., 2006; Michalopoulos, 2011). Among them, epi- dermal growth factor (EGF) has important function in LR. EGF, a multifunctional growth factor, is known as a regulator in a wide variety of physiological processes
higher mortality rate in newborn animals relative to wean- ing animals (P = 0.003). Despite this fact, the creation of a newborn experimental model of hepatectomy and liverregeneration in rats (age 5-7 days) is important because these animals correspond to children weighing less than 5 kg. With the development and refinement of surgical tech- niques and microsurgical anastomoses, a series of liver transplants in such babies have been described and per- formed in centers throughout the world. 14,15 As a result, we
Objective: to evaluate whether colectomy, associated with 70% hepatectomy, influences liverregeneration in rats. Methods: we dis- tributed 18 Wistar rats in three groups of six animals each. In group I (sham), we performed laparotomy; In group II, colectomy + 70% hepatectomy; In group III, only 70% hepatectomy. On the 6th postoperative day, we collected blood by cardiac puncture under anesthesia, followed by euthanasia. We performed serum dosages of aspartate aminotransferase (AST), alanine aminotransferase (ALT), albumin and alkaline phosphatase (AF), hepatocyte growth factor (HGF) and transforming growth factor-α (TGF-α). We calculated liverregeneration by the formula: liver weight ratio per 100g body weight at the time of euthanasia / liver weight preoperatively projected for 100g body weight × 100. Results: ALT and AST levels were significantly lower in group II when compared with group III (p<0.001). Albuminemia showed significantly higher levels in group II. Levels of HGF and TGF- in group II were significantly higher than in group III. The percentage of hepatic regeneration was significantly higher in group II than in group III. Conclusion: Colectomy performed simultaneously with 70% hepatectomy had a positive influence on liverregeneration in rats. Further research is needed to reveal the molecular mechanisms of this effect and to characterize the colon influence in liver physiology.
Methods: thirty-six Wistar rats, males, adults were used, weighing between 195 and 330 g assigned to control and groups. The supplementation group received the diet by gavage and were killed after 24h, 72h and seven days. Evaluation of regeneration occurred through analysis of weight gain liver, serum aspartate aminotransferase, alanine aminotransferase, gamma- glutamyltranspeptidase, and mitosis of the liver stained with H&E. Results Results Results Results: the diet supplemented group showed no statistical Results difference (p>0.05) on the evolution of weights. Administration of fatty acids post-hepatectomy had significant reduction in gamma glutamyltransferase levels and may reflect liverregeneration. Referring to mitotic index, it did not differ between period of times among the groups. Conclusion Conclusion Conclusion Conclusion Conclusion: supplementation with fatty acids in rats undergoing 60% hepatic resection showed no significant interference related to liverregeneration.
A significant association of STAT3 activation with late recurrence was suggested by the gene network analysis with the HIR signature and later validated by immunohistostaining of the surrounding liver tissues. Our findings are in good agreement with previous studies demonstrating that STAT3 is necessary for both liverregeneration following hepatic injury and hepatocarcinogen- esis in mouse models [43,45]. The importance of STAT3 in de novo recurrence is supported by the concordant high expression of interleukin 6 (IL6) , a key upstream regulator of STAT3 in the liver. An earlier study demonstrated that IL6 is accountable for a higher incident rate of HCC in males than in females in a mouse model , suggesting that higher IL6 expression and the subsequent activation of STAT3 in surrounding liver ‘‘prime’’ events for tumor development. STAT3 can be also activated by other cytokines such as IL22 and leptin [48,49], suggesting that diverse pathways might be involved in activation of STAT3 and increase susceptibility to de novo recurrence. The concomitant activation of NOTCH1, and its potential cross-talk with STAT3 in the HIR subgroup, is also intriguing because of the roles of both proteins in stem cells and organ regeneration . Previous studies also support potential roles of other identified genes in the prediction model. For example, higher expression of RALGDS in HCC was previously reported, and silencing or inhibition of RALGDS significantly reduced tumorigenesis in an animal model . Higher expression of CCL20 and SOCS3 was significantly associated with poor prognosis after curative resection of HCC tumors [52,53]. Our data provide evidence for the current notion postulating that micrometastasis and de novo development are accountable for early and late recurrence, respectively, and suggest an update to the notion in recognition of the distinct molecular mechanisms for each type of recurrence that are not shared.
The issues of cities development trajectories and culture-led local regeneration have become more and more present within the urban studies (see Miles and Paddison, 2005, for a critical review). These emerging phenomena arose in a period of turbulent socio- economic change fuelled by globalization processes. According to these dynamics, virtually all of the cities, that are characterized by advanced levels of industrial development, are experiencing a massive re-shaping of their productive sectors. In this context, culture and creativity are clear examples of sectors characterized by high value added, high incidence of service and other intangible components, and by an interesting socio-economic impact on the local economies. At the European level this issue finds its institutional collocation inside the European research strategies developed by KEA Europe Affair. The year 2009 was designated as the “European Year of Creativity and Innovation”. In that sense the European Commission (E.C., 2009) has drawn attention to the usefulness of culture and creativity in sustaining the emergence of innovation-led economy within the EU integration policies 741 .
The germplasm used in the present study represented various types of Brazilian cultivars. Although these cultivars performed differently upon in vitro culture conditions, the regeneration system reported here is efficient, reliable, and cultivar-independent. Based on previous research using onion, shallot and garlic calli as explants for transformation (Zheng et al., 2001, 2003, 2004), root segment calli from both apical and non-apical tissues are an appropriate starting point for garlic genetic transformation via Table 2. Indirect in vitro organogenesis regeneration of eight marketable garlic cultivars.
Gastrinomas are generally localized in pancreas, duodenum and lymphonodes, within the so called “gastrinoma’s triangle” . In 5% of the cases, it may arise from liver, stomach, ovarium, kidneys, parathyroid, omentum, jejunum and heart. We describe a case of a fifteen-year-old boy with a primary gastrinoma of the liver, treated by right-hepatectomy.