Mesenchymal stem cell-derived exosomes promote hepatic regeneration in drug-induced liver injury models
© Tan et al.; licensee BioMed Central Ltd. 2014
Received: 21 February 2014
Accepted: 2 June 2014
Published: 10 June 2014
Mesenchymal stem cell-conditioned medium (MSC-CM) has been shown to have protective effects against various cellular-injury models. This mechanism of protection, however, has yet to be elucidated. Recently, exosomes were identified as the active component in MSC-CM. The aim of this study is to investigate the effect of MSC-derived exosomes in an established carbon tetrachloride (CCl4)-induced liver injury mouse model. This potential effect is then validated by using in vitro xenobiotic-induced liver-injury assays: (1) acetaminophen (APAP)- and (2) hydrogen peroxide (H2O2)-induced liver injury.
The exosomes were introduced concurrent with CCl4 into a mouse model through different routes of administration. Biochemical analysis was performed based on the blood and liver tissues. Subsequently the exosomes were treated in APAP and H2O2-toxicants with in vitro models. Cell viability was measured, and biomarkers indicative of regenerative and oxidative biochemical responses were determined to probe the mechanism of any hepatoprotective activity observed.
In contrast to mice treated with phosphate-buffered saline, CCl4 injury in mice was attenuated by concurrent-treatment exosomes, and characterized by an increase in hepatocyte proliferation, as demonstrated with proliferating cell nuclear antigen (PCNA) elevation. Significantly higher cell viability was demonstrated in the exosomes-treated group compared with the non-exosome-treated group in both injury models. The higher survival rate was associated with upregulation of the priming-phase genes during liver regeneration, which subsequently led to higher expression of proliferation proteins (PCNA and cyclin D1) in the exosomes-treated group. Exosomes also inhibited the APAP- and H2O2-induced hepatocytes apoptosis through upregulation of Bcl-xL protein expression. However, exosomes do not mitigate hepatocyte injury via modulation of oxidative stress.
In summary, these results suggest that MSC-derived exosomes can elicit hepatoprotective effects against toxicants-induced injury, mainly through activation of proliferative and regenerative responses.
Liver is one of the few organs in the body that possesses an immense capacity to regenerate through the replication of mature functioning liver cells . This protective response is of clinical significance particularly during liver injury. However, when the injury progresses into a state of functional impairment, it can overwhelm or even inhibit the intrinsic regenerative potential, leading to severe consequences that include acute liver failure or death . Today, xenobiotic (drugs)-induced liver injury accounts for more than 50% of acute liver failure in the United States and has become a major clinical problem . To overcome this issue, we must explore potential therapeutic agents that can prevent further damage to the injured liver cells. Alternatively, agents that can stimulate remnant liver cells to regenerate could also help to restore liver function in a timely manner and to avert longer-term sequelae.
Mesenchymal stem cell (MSC)-based therapy has been extensively investigated in the area of regenerative medicine for different organs (heart, kidney, lung, and liver) because of its ability to differentiate and transdifferentiate into various tissue types (for example, from osteoblasts into adipocytes and chondrocytes) , stimulate regeneration, and repair damaged tissue/organs . However, recent reports showed that the bioactive soluble factors found in MSC-conditioned medium (MSC-CM) can stimulate host responses for repair and also reduce the liver-injury level in several different injury models, even without MSC [6–8]. As compared to cell-based therapies, MSC-CM (non-cell-based) therapies are generally preferred because they are less likely to trigger immune response, rendering them safer to use. In addition, they are more amenable to reformulation to support different routes of administration.
Recently, proteomic and computational analysis on the MSC-CM revealed that the secretome in this condition medium (CM) could exert potential modulating effects on tissue repair and regeneration in heart, kidney, and liver . More important, it was demonstrated that the observed cardiac-function preservation was derived from exosomes, a fraction of particle size with 55–100 nm, bi-lipid membrane secreted microvesicle purified from the MSC-CM . However, it remains to be determined if these size-purified exosomes are able to enrich the hepatoregenerative effect found previously in the conditioned medium, and whether they are efficacious in overcoming liver injury. Furthermore, their ability to respond to different mechanisms of liver injury has not been investigated.
Therefore, this work set out first to explore the potential therapeutic effect in an established CCl4-induced acute liver-injury mouse model followed by characterizing this potential effect by using liver cell-culture models of well-defined xenobiotic-induced liver injury.
Materials and methods
Materials and reagents
Dexamethasone, nicotinamide, gentamicin, HEPES, NaCl, EDTA, glycerine, Triton-X, sodium fluoride, sodium orthovanadate, phenylmethanesulfonyl fluoride (PMSF), aprotinin, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), acetaminophen (APAP), hydrogen peroxide (H2O2), carbon tetrachloride (CCl4), and olive oil were obtained from Sigma Chemical (St. Louis, MO, USA). Dulbecco modified Eagle medium/Ham F12 (DMEM/F12), and Superscript III First-strand Synthesis System are products of Invitrogen (Carlsbad, CA, USA). Insulin, transferrin, and selenium (ITS) is from BD (Franklin Lakes, NJ, USA). Dimethylsulfoxide (DMSO) was obtained from Merck (Darmstadt, Germany). SYBR Green PCR master mix was obtained from Applied Biosystems (Warrington, UK). RNeasy mini kit is a product of Qiagen (Hilden, Germany). Phosphate-buffered saline (PBS) and all primers were synthesized by 1st BASE Oligos (Singapore). Primary antibodies were purchased from the following companies: phospho (Tyr705)-STAT 3, proliferating cell nuclear antigen (PCNA), NF-κB p65, cyclin D and cyclin E, Cell Signaling Technology (Danvers, MA, USA); hepatocyte growth factor (HGF), Abcam (Cambridge, UK). MSC-derived exosomes was prepared and purified as described previously .
Animals and diets
Six-week-old male C57BL/6 mice (21 to 27 g) were supplied by the Lab Animals Centre, Singapore. Animals were housed in animal-holding units in National University of Singapore (NUS) at a constant temperature in a 12/12-hour light/dark cycle. All animal procedures were carried out according to a protocol approved by the National University of Singapore Institutional Animal Care & Use Committee, Protocol number 123/09. The mice were acclimatized for 2 weeks before use; the mice were 8 weeks of age at the time of experiments. Before the experiment, mice were allowed free access to food and water.
Preparation of MSC-derived exosomes
The collection of fetal tissue was carried out under a KK Women’s and Children’s Hospital (KKH) IRB-approved protocol (EC200804062) . Only patients who had already consented to Termination of Pregnancy (TOP) in theKKH Outpatient Clinic were recruited. Recruitment was carried out in strict adherence to KKH IRB regulations to ensure the patient’s rights and privacy, and to provide confidential counseling for the patient’s fully informed consent to voluntary donation. The patients gave their informed consent for fetal tissue to be collected and used in this study. The hESC-derived HuES9.E1 MSCs were used for the production of MSC secretion in the form of CM, as described .
To harvest MSC secretion, 80% of the confluent HuES9. E1 cultures were washed with PBS, transferred to a chemically defined, serum-free culture medium for an overnight incubation, washed with PBS, and cultured in fresh, chemically defined, serum-free culture medium for 3 days. The CM was collected, clarified by centrifugation, and was concentrated 50× by tangential flow filtration (TFF) by using a membrane with a 100-kDa MWCO (Sartorius, Goettingen, Germany). CM was fractionated by high-performance liquid chromatography (HPLC) (TSK Guard column SWXL, 6 × 40 mm and TSK gel G4000 SWXL, 7.8 × 300 mm, Tosoh Corp., Tokyo, Japan).
Exosomes were collected from the first peak of the elution and concentrated by using 100 kDa MWCO filter (Sartorius). The identity of the exosomes used in this study had been previously characterized by Lai et al.. Exosomes were filtered with a 0.22-μm filter and stored in a -20°C freezer until use.
CCl4 and exosomes administration
All mice received a single dose of CCl4 (3% vol/vol in olive oil) at 0.05 ml/kg body weight by intraperitoneal injection (i.p.). 0.4 μg (100 μl) exosomes or 100 μl PBS were administered through intrasplenic injection (i.s.) for the treatment group and the vehicle-control group, respectively. All mice were killed 24 hours after CCl4, blood, and livers were collected. The aspartate aminotransferase (AST) and alanine aminotransferase (ALT) activities in the serum were analyzed by using Cobas 4000 analyzer series (Roche, Basel, Switzerland).
Cell lines and culture conditions
Three different types of hepatocytes were used in the study of cell viability. TAMH, an immortalized mouse hepatocyte line derived from transgenic MT42 male mice overexpressing TGF-α , was used as a metabolically competent liver cell line that reproduced features of cytotoxicity to support this investigation, whereas THLE-2 is an immortalized primary human hepatocyte that expresses phenotypic characteristics of normal adult liver epithelial cells .
In contrast with the normal cell line, we also investigated the effect of exosomes in HuH-7, a well-differentiated human hepatocarcinoma cell line. This comparison will help to strengthen the evidence of therapeutic effect of exosomes against different liver-injury model in various liver cell types.
Immortalized murine transforming growth factor alpha (TGF-α) transgenic hepatocyte (TAMH) cells  (obtained as a kind gift from Prof Nelson Fausto, University of Washington, Seattle, WA, USA), were maintained in DMEM/F12 supplemented with 5 mg/ml insulin, 5 mg/ml transferrin, 5 ng/ml selenium, 100 nM dexamethasone, 10 mM nicotinamide, and 0.01% (vol/vol) gentamicin. HuH-7 obtained from HSRRB (Tokyo, Japan), a human hepatocarcinoma cell line was grown in Dulbecco modified Eagle medium (DMEM) (Sigma) supplemented with 10% fetal bovine serum (FBS) (Hyclone). THLE-2 obtained from ATCC (Manassas, VA, USA), a human normal liver cell, was cultured in LHC-9 medium supplemented with 10% FBS (Hyclone). The flasks used were pre-coated with a mixture of 0.01 mg/ml fibronectin, 0.03 mg/ml bovine collagen type I, and 0.1 mg/ml bovine serum albumin (BSA) dissolved in LHC-9 basal medium.
All cells were maintained at 37°C in humidified 95% air and 5% CO2 atmosphere, passaged and seeded at 80% to 90% confluency.
In vitro cell-viability assay
With 96-well plates, TAMH cells were seeded at a density of 6 × 103 cells/well in 200 μl DMEM/F12 medium, whereas THLE-2 cells were seeded at a density of 1 × 104 cells/well in 200 μl LHC-9 medium overnight. HuH-7 cells were seeded at a density of 5 × 103 cells/well for 72-hour treatment, and 7.5 × 103 cells/well for 24-hour treatment in 200 μl DMEM medium overnight. TAMH, HuH-7, and THLE-2 cells were treated with various concentrations of exosomes in their respective culturing medium for 24 or 72 hours. Cell-viability assays were performed after the respective incubation time to determine the cytotoxicity of exosomes. In TAMH cells, 2 mM APAP or 350 μM H2O2 was treated in the TAMH cells concurrent with 0.05 μg/ml or 0.1 μg/ml of exosomes for 24 hours before performing the cell-viability test. HuH-7 and THLE-2 cells were treated with 5 mM and 3.5 mM APAP, respectively, with 0.05 μg/ml or 0.1 μg/ml of exosomes for 72 hours before performing the cell-viability test.
PBS was used as background for exosomes treated in the control groups. After incubation of toxicants with different concentrations of exosomes, the cell viability was evaluated with MTT assay. Then 20 ml of 5 mg/ml MTT was dissolved in PBS. After the incubation period, the media was aspirated, and the formazan crystals in cells were dissolved in 200 μl of DMSO and 25 μl of Sorenson buffer . The absorbance was read at 570 nm by using a Tecan microtiter plate reader. Cell-viability percentage was expressed as a ratio of cells exposed to different concentrations of toxicants with those of vehicle controls.
Reverse transcription and quantitative real-time polymerase chain reaction
Sequences of primers used in real-time PCR reaction
Forward primer (5ʹ- > 3ʹ)
Reverse primer (5ʹ- > 3ʹ)
In vitro hepatocytes (TAMH cells)/In vivo liver (mouse)
ATG AGC ACA GAA AGC ATG ATC
TAC AGG CTT GTA ACT CGA ATT
AGT TGC CTT CTT GGG ACT GA
TCC ACG ATT TCC CAG AGA AC
CAC CTT GGA GTT CAC CCA GT
ACC ACT CGT ACT TGG GAT GC
CTC CCT GAA GCC GTA CAC AT
GCT CGG CTT CCAA GTA TTG AG
AAG TTT GCC TTG ACC CTG AA
AGG CAC ATC AGG TAC GAT CC
CAG GTG ATG CTG ACA GAG GA
ATG GCA TAA ATT CCC ACT GC
CCG GCT ACA ACG TCA AGT TT
CGG CAG GTC CTT CTC TAT CA
ACC ACG AGG AAG ACG AAA TG
GGT GAC CAG CTC CTC TGA AG
GGC CAA GGG AGA TGT TAC AA
CCT TGG ACT CCC ACA GAC AT
GGC ATT GCT CTC AAT GGAC AA
CCG AGGT TGG GAT AGG GCC
Mouse liver tissue was homogenized by using glass dounce homogenizer. Proteins were lysated with 800 μl of lysis buffer containing 50 mM HEPES pH 7.5, 150 mM NaCl, 1 mM EDTA, 10% glycerine, 1% triton-X, 0.5 M sodium fluoride, 100 mM sodium orthovanadate, 100 mM PMSF, and 20 mg/ml aprotinin. TAMH cells were seeded at 2 × 106 in a 10-cm dish with DMEM/F12 medium overnight. The medium was changed to DMEM/F12 low glucose and cultured for 2 days. Cells were then treated with respective concentrations of APAP or H2O2 and exosomes for 24 hours. All cells were harvested for Western blot analysis. Cell pellets were lysed with 100 μl of RIPA lysis buffer containing 50 mM Tris, 150 mM NaCl, 0.1% SDS, 0.5% sodium deoxycholate, 1% NP-40, 0.5 M sodium fluoride, 100 mM sodium orthovanadate, 100 mM PMSF, and 20 mg/ml aprotinin. The 40 mg total protein per mouse sample and 10 μg per cell sample were resolved in 10% SDS-PAGE and immobilized on PVDF membrane. The membranes were blocked in 5% milk for 1 hour and incubated in respective primary antibodies overnight (pSTAT3, 1:1,000; NF-κB, 1:2,000; Bcl-xL 1:1,000; PCNA, 1:10,000; and cyclin D1, 1:1,000), followed by respective secondary antibody (1:10,000) incubation for 1 hour. All membranes were visualized by using chemiluminescence substrate (Pierce Biotechnology, Rockford, IL, USA). The band intensities were normalized against actin and were quantified by using ImageJ software.
Data were expressed as means ± standard error of means (SEM) and analyzed by using Student two-tailed t test. Statistical significance of the difference was accepted at P-values of less than 0.05.
Exosomes reduced liver-injury level in ALT and AST after CCl4 treatment in vivo
Exosomes induced hepatocyte-regenerative genes expression in liver tissue after CCl4-induced injury
Exosomes exerted no intrinsic cytotoxicity while demonstrating increased cell viability in TAMH, THLE-2, and Huh-7 hepatocytes after APAP- or H2O2-induced injuries
Exosomes regulated hepatocytes proliferation through inducing quiescent hepatocytes (G0) to re-enter the cell cycle (G1) at the priming phase after APAP- or H2O2-induced injury
In the APAP-injury model, MIP-2 demonstrated the highest elevation with 3.7-fold, followed by 2.4-fold in COX-2 and twofold in iNOS. The elevation trend differed slightly in the H2O2-injury model, whereby the same amount of exosomes was found to increase iNOS expression the most significantly, by approximately 2.9-fold, followed by twofold increase in all three MIP-2, COX-2, and IL-6 genes compared with the non-exosomes-treated group (Figure 4B).
Exosomes induced transcription factors expression during the G1 phase of cell cycle after APAP- or H2O2-induced injury
Exosomes upregulated the expression of cell-proliferation markers (at G1 and S phase) after APAP- or H2O2-induced injury
Exosomes protected hepatocytes from apoptosis by decreasing caspase 3/7 level while upregulating antiapoptotic gene Bcl- xL
Exosomes did not regulate the antiapoptotic effect through antioxidative genes
We hypothesized that the size-purified subset of exosomes derived from MSC-CM can play a role in mitigating xenobiotic-induced liver injury. As a proof of concept, we explored such effect by using classic models of liver toxicity (that is, CCl4-induced hepatic injury in mouse). We further considered the selectivity and the mechanism of the anticipated protection by using in vitro models of well-defined mechanisms of toxicity: APAP and H2O2. Accordingly, our experiments demonstrated that the MSC-derived exosomes accelerated in vivo liver regeneration in CCl4-injured mice. In addition, we found that exosomes consistently caused increased cell viability after the injury arising from both APAP- and H2O2.
In an in vivo study, CCl4 was used as a classic and well-established model of acute liver injury. The administration of MSC-derived exosomes was found to reverse CCl4-induced injury in mice with active proliferation of hepatocytes, most clearly indicated with the expression of PCNA, cyclin D1, and cyclin E. These proliferation effects were in line with the in vitro findings in which exosomes were able to sustain higher cell viability of hepatocytes after injuries cytotoxic doses of both APAP and H2O2. We qualified the regenerative potential of these exosomes by investigating the biochemical machinery behind the effect, beginning from the priming phase that triggers growth-arrested injured cells to reenter cell cycle.
IL-6  and TNF-α  are prominent priming factors in initiating liver regeneration on injury by committing cells to cell cycling. Yamada et al.  reported defective DNA synthesis in TNFR-I-deficient mice after partial hepatectomy, whereas IL-6 injection restored the liver regeneration. Besides these, iNOS has also been demonstrated to mediate the formation of nitric oxide (NO) [23, 24] and COX-2, which catalyzes the rate-limiting step in the synthesis of prostaglandins (PGs) that control the regenerative process. MIP-2 was also found to enhance hepatocytes proliferation through upregulating nuclear translocation of STAT-3 during the process of recovery from injury [25–27]. Our results demonstrated the increase in expressions of all these priming-phase factors after exosomes treatment alongside improved cell viability. Despite the use of a monoculture system comprising exclusively hepatocytes, we were able to pick up these cytokine perturbations, consistent with other similar liver cell-line studies reported in the literature [28, 29].
TNF initiates liver regeneration by activating the NF-κB and IL-6-dependent pathway, which involves STAT3 transcription factor  in the G1 phase. In the current investigation, Western blot demonstrated that injured hepatocytes with exosomes treatment markedly increased the expression of NF-κB and STAT3 transcription factors. The IL-6/STAT3 pathway subsequently innervates cell-cycle progression from G1 to S phase, as shown in the upregulation of cyclin D1 and PCNA after exosomes treatment. In line with the priming factors and transcription factors, all these upregulated gene and protein expressions collectively support a restoration of overall cell viability, suggesting that the exosomes therapy likely mediates the liver repair from acute liver injury through induction of liver regeneration in the hepatocytes.
In the process of characterizing the mode of action of exosomes, we were aware of a possible differential response to the type of cellular injury invoked by the toxicants. Hence, two mechanistically distinct injury models were investigated. APAP is a well-studied hepatotoxicant that mediates the liver-injury pathway principally through generation of its reactive metabolite, N-acetyl-p-benzoquinoneimine (NAPQI) and subsequent binding to hepatic proteins , whereas H2O2 mediates liver injury solely through the oxidative-stress pathway. However, a common effect observed in both injuries is the manifestation of mitochondrial dysfunction, leading to apoptosis [32, 33]. Because the exosomes demonstrated a protective or injury-repair effect in both models, albeit to a different extent, a possible effect in mitigating oxidative stress was investigated. Previously, STAT3 was also shown to regulate mitochondrial Bcl-xL and MnSOD expression in inhibiting caspase 3 during apoptosis [18, 20]. Our results demonstrated that the Bcl-xL expression increased with higher expression of STAT3 after the treatment of exosomes, whereas caspase 3/7 expression was significantly decreased in the APAP-injury group treated with exosomes. This suggests that exosomes were able to alleviate some mitochondrial dysfunction as part of its plethora of protective mechanisms in the antiapoptosis pathway. However, exosomes did not demonstrate a role in regulating antioxidative genes, and hence did not mitigate injury via modulation of oxidative stress. These suggest that exosomes primarily assert antiapoptotic, proliferative, and regenerative hepatic responses to liver damage.
Follow-up work would be necessary to determine the hepatoprotective components within the exosomes. Exosomes have been shown to contain biologically active proteins, lipids, mRNA, and miRNA [34–36]. Lai et al. profiled a total of 857 proteins in the exosomes by using mass spectrometry and antibody array . Here, few of the prominent proteins that are potential components involved in the liver-regeneration process were identified. Among them, IL6ST/gp130, TNFRSF1A/TNFR1, and CXCL2/MIP-2 proteins were found to be associated with the priming factors during liver regeneration. IL6ST plays an important role in mediating the IL-6/STAT3 pathway, which initiates the hepatocytes protection [38–40]. Likewise, CXCL2/MIP-2 enhances hepatocytes proliferation through upregulating STAT-3 during the process of recovery from injury [25–27]. These proteinaceous contents could be released from the exosomes intrahepatically, triggering the higher expressions of IL-6, TNF-α, and MIP-2 after treatment, as shown in our results.
Apart from the priming factors, HGF and hepatocyte growth factor receptor (HGFR) proteins, which are among the most potent stimulators of hepatocyte growth in liver regeneration [41, 42], can also be found in the exosomes. These proteins might be working synergistically with the priming factors in accelerating and promoting liver regeneration, leading to a restoration of homeostasis. However, it remains to be determined whether miRNAs or other mRNAs found in the exosomes may have any impact on the proliferation effect of the hepatocytes.
In conclusion, exosomes treatment demonstrated liver recovery after several toxicants-induced injuries. It is mediating this effect, likely through maintaining the liver homeostasis, that primarily includes inducing hepatocytes regeneration. This treatment may present a novel adjunctive therapy in drug-induced liver toxicity, in lieu of the limited availability of the conventional liver transplants. Its attractiveness also lies largely on its non-cell-based system, which reduces the risk of tissue incompatibility, while using an abundant and reproducible supply. This work supports further investigation to clarify the exact mechanism of action so that the intended therapeutic effect can be translated and optimized.
bovine serum albumin
Dulbecco modified Eagle medium/Ham F12
fetal bovine serum
glutathione peroxidase 4
hematoxylin and eosin
hepatocyte growth factor
hepatocyte growth factor receptor
high-performance liquid chromatography
inducible nitric oxide synthase
insulin, transferrin, selenium
macrophage inflammatory protein 2
manganese superoxide dismutase
mesenchymal stem cell-conditioned medium
proliferating cell nuclear antigen
quantitative real-time polymerase chain reaction
tangential flow filtration
transforming growth factor alpha
tumor necrosis factor alpha.
This work was supported by NMRC grant R-148-000-125-275 (HKH) and BMRC intramural research funds (SKL). The authors thank A/Prof Mei Lin Go for her mentorship and guidance.
- Thorgeirsson SS: Hepatic stem cells in liver regeneration. FASEB J. 1996, 10: 1249-1256.PubMedGoogle Scholar
- Navarro VJ, Senior JR: Drug-related hepatotoxicity. N Engl J Med. 2006, 354: 731-739.View ArticlePubMedGoogle Scholar
- Daniels D, Grytdal S, Wasley A: Centers for Disease Control and Prevention: surveillance for acute viral hepatitis, United States, 2007. MMWR Surveill Summ. 2009, 58: 1-27.PubMedGoogle Scholar
- Song L, Tuan RS: Transdifferentiation potential of human mesenchymal stem cells derived from bone marrow. FASEB J. 2004, 18: 980-982.PubMedGoogle Scholar
- Le Blanc K, Pittenger M: Mesenchymal stem cells: progress toward promise. Cytotherapy. 2005, 7: 36-45.View ArticlePubMedGoogle Scholar
- Zagoura DS, Roubelakis MG, Bitsika V, Trohatou O, Pappa KI, Kapelouzou A, Antsaklis A, Anagnou NP: Therapeutic potential of a distinct population of human amniotic fluid mesenchymal stem cells and their secreted molecules in mice with acute hepatic failure. Gut. 2012, 61: 894-906.View ArticlePubMedGoogle Scholar
- Kanazawa H, Fujimoto Y, Teratani T, Iwasaki J, Kasahara N, Negishi K, Tsuruyama T, Uemoto S, Kobayashi E: Bone marrow-derived mesenchymal stem cells ameliorate hepatic ischemia reperfusion injury in a rat model. PLoS One. 2011, 6: e19195-PubMed CentralView ArticlePubMedGoogle Scholar
- Yan YM, Xu WR, Qian H, Si Y, Zhu W, Cao HL, Zhou HX, Mao F: Mesenchymal stem cells from human umbilical cords ameliorate mouse hepatic injury in vivo. Liver Int. 2009, 29: 356-365.View ArticlePubMedGoogle Scholar
- Sze SK, de Kleijn DP, Lai RC, Khia Way Tan E, Zhao H, Yeo KS, Low TY, Lian Q, Lee CN, Mitchell W, El Oakley RM, Lim SK: Elucidating the secretion proteome of human embryonic stem cell-derived mesenchymal stem cells. Mol Cell Proteomics. 2007, 6: 1680-1689.View ArticlePubMedGoogle Scholar
- Lai RC, Arslan F, Lee MM, Sze NS, Choo A, Chen TS, Salto-Tellez M, Timmers L, Lee CN, El Oakley RM, Pasterkamp G, de Kleijn DP, Lim SK: Exosome secreted by MSC reduces myocardial ischemia/reperfusion injury. Stem Cell Res. 2010, 4: 214-222.View ArticlePubMedGoogle Scholar
- Lai RC, Arslan F, Tan SS, Tan B, Choo A, Lee MM, Chen TS, Teh BJ, Eng JK, Sidik H, Tanavde V, Hwang WS, Lee CN, El Oakley RM, Pasterkamp G, de Kleijn DP, Tan KH, Lim SK: Derivation and characterization of human fetal MSCs: an alternative cell source for large-scale production of cardioprotective microparticles. J Mol Cell Cardiol. 2010, 48: 1215-1224.View ArticlePubMedGoogle Scholar
- Wu JC, Merlino G, Cveklova K, Mosinger B, Fausto N: Autonomous growth in serum-free medium and production of hepatocellular carcinomas by differentiated hepatocyte lines that overexpress transforming growth factor alpha 1. Cancer Res. 1994, 54: 5964-5973.PubMedGoogle Scholar
- Pfeifer AM, Cole KE, Smoot DT, Weston A, Groopman JD, Shields PG, Vignaud JM, Juillerat M, Lipsky MM, Trump BF: Simian virus 40 large tumor antigen-immortalized normal human liver epithelial cells express hepatocyte characteristics and metabolize chemical carcinogens. Proc Natl Acad Sci U S A. 1993, 90: 5123-5127.PubMed CentralView ArticlePubMedGoogle Scholar
- Coe KJ, Jia Y, Ho HK, Rademacher P, Bammler TK, Beyer RP, Farin FM, Woodke L, Plymate SR, Fausto N, Nelson SD: Comparison of the cytotoxicity of the nitroaromatic drug flutamide to its cyano analogue in the hepatocyte cell line TAMH: evidence for complex I inhibition and mitochondrial dysfunction using toxicogenomic screening. Chem Res Toxicol. 2007, 20: 1277-1290.PubMed CentralView ArticlePubMedGoogle Scholar
- Plumb JA, Milroy R, Kaye SB: Effects of the pH dependence of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide-formazan absorption on chemosensitivity determined by a novel tetrazolium-based assay. Cancer Res. 1989, 49: 4435-4440.PubMedGoogle Scholar
- Reid AB, Kurten RC, McCullough SS, Brock RW, Hinson JA: Mechanisms of acetaminophen-induced hepatotoxicity: role of oxidative stress and mitochondrial permeability transition in freshly isolated mouse hepatocytes. J Pharmacol Exp Ther. 2005, 312: 509-516.View ArticlePubMedGoogle Scholar
- Kirillova I, Chaisson M, Fausto N: Tumor necrosis factor induces DNA replication in hepatic cells through nuclear factor kappaB activation. Cell Growth Differ. 1999, 10: 819-828.PubMedGoogle Scholar
- Fukada T, Hibi M, Yamanaka Y, Takahashi-Tezuka M, Fujitani Y, Yamaguchi T, Nakajima K, Hirano T: Two signals are necessary for cell proliferation induced by a cytokine receptor gp130: involvement of STAT3 in anti-apoptosis. Immunity. 1996, 5: 449-460.View ArticlePubMedGoogle Scholar
- Haga S, Terui K, Zhang HQ, Enosawa S, Ogawa W, Inoue H, Okuyama T, Takeda K, Akira S, Ogino T, Irani K, Ozaki M: Stat3 protects against Fas-induced liver injury by redox-dependent and -independent mechanisms. J Clin Invest. 2003, 112: 989-998.PubMed CentralView ArticlePubMedGoogle Scholar
- Terui K, Enosawa S, Haga S, Zhang HQ, Kuroda H, Kouchi K, Matsunaga T, Yoshida H, Engelhardt JF, Irani K, Ohnuma N, Ozaki M: Stat3 confers resistance against hypoxia/reoxygenation-induced oxidative injury in hepatocytes through upregulation of Mn-SOD. J Hepatol. 2004, 41: 957-965.View ArticlePubMedGoogle Scholar
- Clavien PA: IL-6, a key cytokine in liver regeneration. Hepatology. 1997, 25: 1294-1296.View ArticlePubMedGoogle Scholar
- Yamada Y, Kirillova I, Peschon JJ, Fausto N: Initiation of liver growth by tumor necrosis factor: deficient liver regeneration in mice lacking type I tumor necrosis factor receptor. Proc Natl Acad Sci U S A. 1997, 94: 1441-1446.PubMed CentralView ArticlePubMedGoogle Scholar
- Hortelano S, Dewez B, Genaro AM, Diaz-Guerra MJ, Bosca L: Nitric oxide is released in regenerating liver after partial hepatectomy. Hepatology. 1995, 21: 776-786.PubMedGoogle Scholar
- Fausto N, Campbell JS, Riehle KJ: Liver regeneration. Hepatology. 2006, 43: S45-S53.View ArticlePubMedGoogle Scholar
- Ren X, Carpenter A, Hogaboam C, Colletti L: Mitogenic properties of endogenous and pharmacological doses of macrophage inflammatory protein-2 after 70% hepatectomy in the mouse. Am J Pathol. 2003, 163: 563-570.PubMed CentralView ArticlePubMedGoogle Scholar
- Colletti LM, Green M, Burdick MD, Kunkel SL, Strieter RM: Proliferative effects of CXC chemokines in rat hepatocytes in vitro and in vivo. Shock. 1998, 10: 248-257.View ArticlePubMedGoogle Scholar
- Clarke CN, Kuboki S, Tevar A, Lentsch AB, Edwards M: CXC chemokines play a critical role in liver injury, recovery, and regeneration. Am J Surg. 2009, 198: 415-419.PubMed CentralView ArticlePubMedGoogle Scholar
- Dong W, Simeonova PP, Gallucci R, Matheson J, Fannin R, Montuschi P, Flood L, Luster MI: Cytokine expression in hepatocytes: role of oxidant stress. J Interferon Cytokine Res. 1998, 18: 629-638.View ArticlePubMedGoogle Scholar
- Rowell DL, Eckmann L, Dwinell MB, Carpenter SP, Raucy JL, Yang SK, Kagnoff MF: Human hepatocytes express an array of proinflammatory cytokines after agonist stimulation or bacterial invasion. Am J Physiol. 1997, 273: G322-G332.PubMedGoogle Scholar
- Gilgenkrantz H, Collin de l'Hortet A: New insights into liver regeneration. Clin Res Hepatol Gastroenterol. 2011, 35: 623-629.View ArticlePubMedGoogle Scholar
- Cohen SD, Pumford NR, Khairallah EA, Boekelheide K, Pohl LR, Amouzadeh HR, Hinson JA: Selective protein covalent binding and target organ toxicity. Toxicol Appl Pharmacol. 1997, 143: 1-12.View ArticlePubMedGoogle Scholar
- Li JM, Zhou H, Cai Q, Xiao GX: Role of mitochondrial dysfunction in hydrogen peroxide-induced apoptosis of intestinal epithelial cells. World J Gastroenterol. 2003, 9: 562-567.PubMed CentralView ArticlePubMedGoogle Scholar
- Meyers LL, Beierschmitt WP, Khairallah EA, Cohen SD: Acetaminophen-induced inhibition of hepatic mitochondrial respiration in mice. Toxicol Appl Pharmacol. 1988, 93: 378-387.View ArticlePubMedGoogle Scholar
- Cocucci E, Racchetti G, Meldolesi J: Shedding microvesicles: artefacts no more. Trends Cell Biol. 2009, 19: 43-51.View ArticlePubMedGoogle Scholar
- Gyorgy B, Szabo TG, Pasztoi M, Pal Z, Misjak P, Aradi B, Laszlo V, Pallinger E, Pap E, Kittel A, Nagy G, Falus A, Buzás EI: Membrane vesicles, current state-of-the-art: emerging role of extracellular vesicles. Cell Mol Life Sci. 2011, 68: 2667-2688.PubMed CentralView ArticlePubMedGoogle Scholar
- Valadi H, Ekstrom K, Bossios A, Sjostrand M, Lee JJ, Lotvall JO: Exosome-mediated transfer of mRNAs and microRNAs is a novel mechanism of genetic exchange between cells. Nat Cell Biol. 2007, 9: 654-659.View ArticlePubMedGoogle Scholar
- Lai RC, Tan SS, Teh BJ, Sze SK, Arslan F, de Kleijn DP, Choo A, Lim SK: Proteolytic potential of the MSC exosome proteome: implications for an exosome-mediated delivery of therapeutic proteasome. Int J Proteomics. 2012, 2012: 971907-PubMed CentralView ArticlePubMedGoogle Scholar
- Klein C, Wustefeld T, Assmus U, Roskams T, Rose-John S, Muller M, Manns MP, Ernst M, Trautwein C: The IL-6-gp130-STAT3 pathway in hepatocytes triggers liver protection in T cell-mediated liver injury. J Clin Invest. 2005, 115: 860-869.PubMed CentralView ArticlePubMedGoogle Scholar
- Dierssen U, Beraza N, Lutz HH, Liedtke C, Ernst M, Wasmuth HE, Trautwein C: Molecular dissection of gp130-dependent pathways in hepatocytes during liver regeneration. J Biol Chem. 2008, 283: 9886-9895.View ArticlePubMedGoogle Scholar
- Wuestefeld T, Klein C, Streetz KL, Betz U, Lauber J, Buer J, Manns MP, Muller W, Trautwein C: Interleukin-6/glycoprotein 130-dependent pathways are protective during liver regeneration. J Biol Chem. 2003, 278: 11281-11288.View ArticlePubMedGoogle Scholar
- Ishii T, Sato M, Sudo K, Suzuki M, Nakai H, Hishida T, Niwa T, Umezu K, Yuasa S: Hepatocyte growth factor stimulates liver regeneration and elevates blood protein level in normal and partially hepatectomized rats. J Biochem. 1995, 117: 1105-1112.PubMedGoogle Scholar
- Jiang WG, Hallett MB, Puntis MC: Hepatocyte growth factor/scatter factor, liver regeneration and cancer metastasis. Br J Surg. 1993, 80: 1368-1373.View ArticlePubMedGoogle Scholar
This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.