Mesenchymal stem cells: A promising therapeutic avenue for nonalcoholic fatty liver disease

Non-alcoholic fatty liver disease(NAFLD)is a pressing global health concern that is associated with metabolic syndrome and obesity.On the basis of the insights provided by Jiang et al,this editorial presents an exploration of the potential of mesenchymal stem cells(MSCs)for NAFLD treatment.MSCs have numerous desirable characteristics,including immunomodulation,anti-inflammatory pro-perties,and tissue regeneration promotion,rendering them attractive candidates for NAFLD treatment.Recent preclinical and early clinical studies have high-lighted the efficacy of MSCs in improving liver function and reducing disease severity in NAFLD models.However,MSC heterogeneity,long-term safety concerns,and unoptimized therapeutic protocols remain substantial challenges.Addressing these challenges through standardized protocols and rigorous clinical trials is essential to the safe and successful application of MSCs in NAFLD mana-gement.Continued research into MSC mechanisms and therapeutic optimization is required to improve treatments for NAFLD and related liver diseases.

Innovative mesenchymal stem cell treatments for fatty liver disease

The incidence of non-alcoholic fatty liver disease(NAFLD)and alcohol-associated liver disease(ALD)is increasing year by year due to changes in the contemporary environment and dietary structure,and is an important public health problem worldwide.There is an urgent need to continuously improve the understanding of their disease mechanisms and develop novel therapeutic strategies.Mesenchymal stem cells(MSCs)have shown promise as a potential therapeutic strategy in therapeutic studies of NAFLD and ALD.NAFLD and ALD have different triggers and their specific mechanisms of disease progression are different,but both involve disease processes such as hepatocellular steatosis and potential fibrosis,cirrhosis,and even hepatocellular carcinoma.MSCs have metabolic regulatory,anti-apoptotic,antioxidant,and immunomodulatory effects that together promote liver injury repair and functional recovery,and have demonstrated positive results in preclinical studies.This editorial is a continuum of Jiang et al’s review focusing on the advantages and limitations of MSCs and their derivatives as therapeutics for NAFLD and ALD.They detail how MSCs attenuate the progression of NAFLD by modulating molecular pathways involved in glucolipid metabolism,inflammation,oxidative stress,endoplasmic reticulum stress,and fibrosis.Based on recent advances,we discuss MSCs and their derivatives as therapeutic strategies for NAFLD and ALD,providing useful information for their clinical application.

Current perspectives on mesenchymal stem cells as a potential therapeutic strategy for non-alcoholic fatty liver disease

Non-alcoholic fatty liver disease(NAFLD)has emerged as a significant health challenge,characterized by its widespread prevalence,intricate natural progression and multifaceted pathogenesis.Although NAFLD initially presents as benign fat accumulation,it may progress to steatosis,non-alcoholic steatohepatitis,cirrhosis,and hepatocellular carcinoma.Mesenchymal stem cells(MSCs)are recognized for their intrinsic self-renewal,superior biocompatibility,and minimal immunogenicity,positioning them as a therapeutic innovation for liver diseases.Therefore,this review aims to elucidate the potential roles of MSCs in alleviating the progression of NAFLD by alteration of underlying molecular pathways,including glycolipid metabolism,inflammation,oxidative stress,endoplasmic reticulum stress,and fibrosis.The insights are expected to provide further understanding of the potential of MSCs in NAFLD therapeutics,and support the development of MSC-based therapy in the treatment of NAFLD.

Mechanism of mesenchymal stem cells in liver regeneration:Insights and future directions

Mesenchymal stem cells(MSCs)are a prevalent source for stem cell therapy and play a crucial role in modulating both innate and adaptive immune responses.Non-alcoholic fatty liver disease(NAFLD)is characterized by the accumulation of triglycerides in liver cells and involves immune system activation,leading to histological changes,tissue damage,and clinical symptoms.A recent publication by Jiang et al,highlighted the potential of MSCs to mitigate in NAFLD progression by targeting various molecular pathways,including glycolipid metabolism,inflammation,oxidative stress,endoplasmic reticulum stress,and fibrosis.In this editorial,we comment on their research and discuss the efficacy of MSC therapy in treating NAFLD.

Enhancing the functionality of mesenchymal stem cells:An attractive treatment strategy for metabolic dysfunction-associated steatotic liver disease?

The intrinsic heterogeneity of metabolic dysfunction-associated fatty liver disease(MASLD)and the intricate pathogenesis have impeded the advancement and clinical implementation of therapeutic interventions,underscoring the critical demand for novel treatments.A recent publication by Li et al proposes mesenchymal stem cells as promising effectors for the treatment of MASLD.This editorial is a continuum of the article published by Jiang et al which focuses on the significance of strategies to enhance the functionality of mesenchymal stem cells to improve efficacy in curing MASLD,including physical pretreatment,drug or chemical pretreatment,pretreatment with bioactive substances,and genetic engineering.

Acute liver failure:A systematic review and network meta-analysis of optimal type of stem cells in animal models

BACKGROUND The therapeutic effects of various stem cells in acute liver failure(ALF)have been demonstrated in preclinical studies.However,the specific type of stem cells with the highest therapeutic potential has not been determined.AIM To validate the efficacy of stem cells in ALF model and to identify the most promising stem cells.METHODS A search was conducted on the PubMed,Web of Science,Embase,Scopus,and Cochrane databases from inception to May 3,2022,and updated on November 16,2022 to identify relevant studies.Two independent reviewers performed the literature search,identification,screening,quality assessment,and data extraction.RESULTS A total of 89 animal studies were included in the analysis.The results of traditional meta-analysis showed that stem cell therapy could significantly reduce the serum levels of alanine aminotransferase[weighted mean difference(WMD)=-181.05(-191.71,-170.39)],aspartate aminotransferase[WMD=-309.04(-328.45,-289.63)],tumor necrosis factor-alpha[WMD=-8.75(-9.93,-7.56)],and interleukin-6[WMD=-10.43(-12.11,-8.76)]in animal models of ALF.Further subgroup analysis and network meta-analysis showed that although mesenchymal stem cells are the current research hotspot,the effect of liver stem cells(LSCs)on improving liver function is significantly better than that of the other five types of stem cells.In addition,the ranking results showed that the possibility of LSCs improving liver function ranked first.This fully proves the great therapeutic potential of LSCs,which needs to be paid more attention in the future.CONCLUSION LSCs may have a higher therapeutic potential.Further high-quality animal experiments are needed to explore the most effective stem cells for ALF.

Targeting the mechano-microenvironment and liver cancer stem cells:a promising therapeutic strategy for liver cancer

Over the past 2 decades,cancer stem cells(CSCs)have been identified as the root cause of cancer occurrence,progression,chemoradioresistance,recurrence,and metastasis.Targeting CSCs is a novel therapeutic strategy for cancer management and treatment.Liver cancer(LC)is a malignant disease that can endanger human health.Studies are increasingly suggesting that changes in the liver mechanical microenvironment are a primary driver triggering the occurrence and development of liver cancer.In this review,we summarize current understanding of the roles of the liver mechano-microenvironment and liver cancer stem cells(LCSCs)in liver cancer progression.We also discuss the relationship between the mechanical heterogeneity of liver cancer tissues and LCSC recruitment and metastasis.Finally,we highlight potential mechanosensitive molecules in LCSCs and mechanotherapy in liver cancer.Understanding the roles and regulatory mechanisms of the mechano-microenvironment and LCSCs may provide fundamental insights into liver cancer progression and aid in further development of novel therapeutic strategies.

The combined application of stem cells and three-dimensional bioprinting scaffolds for the repair of spinal cord injury

Spinal cord injury is considered one of the most difficult injuries to repair and has one of the worst prognoses for injuries to the nervous system.Following surgery,the poor regenerative capacity of nerve cells and the generation of new scars can make it very difficult for the impaired nervous system to restore its neural functionality.Traditional treatments can only alleviate secondary injuries but cannot fundamentally repair the spinal cord.Consequently,there is a critical need to develop new treatments to promote functional repair after spinal cord injury.Over recent years,there have been seve ral developments in the use of stem cell therapy for the treatment of spinal cord injury.Alongside significant developments in the field of tissue engineering,three-dimensional bioprinting technology has become a hot research topic due to its ability to accurately print complex structures.This led to the loading of three-dimensional bioprinting scaffolds which provided precise cell localization.These three-dimensional bioprinting scaffolds co uld repair damaged neural circuits and had the potential to repair the damaged spinal cord.In this review,we discuss the mechanisms underlying simple stem cell therapy,the application of different types of stem cells for the treatment of spinal cord injury,and the different manufa cturing methods for three-dimensional bioprinting scaffolds.In particular,we focus on the development of three-dimensional bioprinting scaffolds for the treatment of spinal cord injury.

Establishing delivery route-dependent safety and efficacy of living biodrug mesenchymal stem cells in heart failure patients

BACKGROUND Mesenchymal stem cells(MSCs)as living biopharmaceuticals with unique properties,i.e.,stemness,viability,phenotypes,paracrine activity,etc.,need to be administered such that they reach the target site,maintaining these properties unchanged and are retained at the injury site to participate in the repair process.Route of delivery(RoD)remains one of the critical determinants of safety and efficacy.This study elucidates the safety and effectiveness of different RoDs of MSC treatment in heart failure(HF)based on phase II randomized clinical trials(RCTs).We hypothesize that the RoD modulates the safety and efficacy of MSCbased therapy and determines the outcome of the intervention.AIM To investigate the effect of RoD of MSCs on safety and efficacy in HF patients.METHODS RCTs were retrieved from six databases.Safety endpoints included mortality and serious adverse events(SAEs),while efficacy outcomes encompassed changes in left ventricular ejection fraction(LVEF),6-minute walk distance(6MWD),and pro-B-type natriuretic peptide(pro-BNP).Subgroup analyses on RoD were performed for all study endpoints.RESULTS Twelve RCTs were included.Overall,MSC therapy demonstrated a significant decrease in mortality[relative risk(RR):0.55,95%confidence interval(95%CI):0.33-0.92,P=0.02]compared to control,while SAE outcomes showed no significant difference(RR:0.84,95%CI:0.66-1.05,P=0.11).RoD subgroup analysis revealed a significant difference in SAE among the transendocardial(TESI)injection subgroup(RR=0.71,95%CI:0.54-0.95,P=0.04).The pooled weighted mean difference(WMD)demonstrated an overall significant improvement of LVEF by 2.44%(WMD:2.44%,95%CI:0.80-4.29,P value≤0.001),with only intracoronary(IC)subgroup showing significant improvement(WMD:7.26%,95%CI:5.61-8.92,P≤0.001).Furthermore,the IC delivery route significantly improved 6MWD by 115 m(WMD=114.99 m,95%CI:91.48-138.50),respectively.In biochemical efficacy outcomes,only the IC subgroup showed a significant reduction in pro-BNP by-860.64 pg/mL(WMD:-860.64 pg/Ml,95%CI:-944.02 to-777.26,P=0.001).CONCLUSION Our study concluded that all delivery methods of MSC-based therapy are safe.Despite the overall benefits in efficacy,the TESI and IC routes provided better outcomes than other methods.Larger-scale trials are warranted before implementing MSC-based therapy in routine clinical practice.

Intraportal mesenchymal stem cell transplantation prevents acute liver failure through promoting cell proliferation and inhibiting apoptosis

BACKGROUND： Transplantation of mesenchymal stem cells （MSCs） has been regarded as a potential treatment for acute liver failure （ALF）, but the optimal route was unknown. The present study aimed to explore the most effective MSCs transplantation route in a swine ALF model. METHODS： The swine ALF model induced by intravenous injection of D-Gal was treated by the transplantation of swine MSCs through four routes including intraportal injection （InP group）, hepatic intra-arterial injection （AH group）, peripheral intravenous injection （PV group） and intrahepatic injection （IH group）. The living conditions and survival time were recorded. Blood samples before and after MSCs trans- plantation were collected for the analysis of hepatic function. The histology of liver injury was interpreted and scored in terminal samples. Hepatic apoptosis was detected by TUNEL assay. Apoptosis and proliferation related protein expressions including cleaved caspase-3, survivin, AKT, phospho-AKT （Ser473）, ERK and phospho-ERK （Tyr204） were analyzed by Western blotting. RESULTS： The average survival time of each group was 10.7± 1.6 days （InP）, 6.0±0.9 days （AH）, 4.7±1.4 days （PV）, 4.3± 0.8 days （IH）, respectively, when compared with the average survival time of 3.8±0.8 days in the D-Gal group. The survival rates between the InP group and D-Gal group revealed a statistically significant difference （P〈0.01）. Pathological and biochemical analysis showed that liver damage was the worst in the D-Gal group, while less injury in the InP group. Histopathological scores revealed a significant decrease in the InP group （3.17±1.04, P〈0.01） and AH group （8.17±0.76, P〈0.05） as compared with that in the D-Gal group （11.50±1.32）. The apoptosis rate in the InP group （25.0%±3.4%, P〈0.01） and AH group （40.5%±1.0% , P〈0.05） was lower than that in the D-Gal group （70.6%±8.5%）. The expression of active caspase-3 was inhibited, while the expression of survivin, AKT, phospho- AKT （Ser473）, ERK and phospho-ERK （Tyr204） was elevated in the InP group. CONCLUSIONS： Intraportal injection was superior to other pathways for MSC transplantation. Intraportal MSC trans- plantation could improve liver function, inhibit apoptosis and prolong the survival time of swine with ALE The transplanted MSCs may participate in liver regeneration via promoting cell proliferation and suppressing apoptosis during the initial stage of ALE

Hepatocytic differentiation of mesenchymal stem cells in cocultures with fetal liver cells

AIM： To investigate the hepatocytic differentiation of mesenchymal stem cells （MSCs） in co-cultures with fetal liver cells （FLC） and the possibility to expand differentiated hepatocytic cells. METHODS： MSCs were marked with green fluorescent protein （GFP） by retroviral gene transduction. Clonal marked MSCs were either cultured under liver stimulating conditions using fibronectin-coated culture dishes and medium supplemented with stem cell factor （SCF）, hepatocyte growth factor （HGF）, epidermal growth factor （EGF）, and fibroblast growth factor 4 （FGF-4） alone, or in presence of freshly isolated FLC. Cells in co-cultures were harvested, and GFP＋ or GFP- cells were separated using fluorescence activated cell sorting. Reverse transcription-polymerase chain reaction （RT-PCR） for the liver specific markers cytokeratin-18 （CK-18）, albumin, and alpha-fetoprotein （AFP） was performed in different cell populations. RESULTS- Under the specified culture conditions, rat MSCs co-cultured with FLC expressed albumin, CK-18, and AFP-RNA over two weeks. At wk 3, MSCs lost hepatocytic gene expression, probably due to overgrowth of the cocultured FLC. FLC also showed a stable liver specific gene expression in the co-cultures and a very high growth potential. CONCLUSION： The rat MSCs from bone marrow can differentiate hepatocytic cells in the presence of FLC in vitro and the presence of MSCs in co-cultures also prorides a beneficial environment for expansion and differentiation of FLC.

Bone marrow derived stem cells for the treatment of end-stage liver disease

End-stage disease due to liver cirrhosis is an important cause of death worldwide. Cirrhosis results from progressive, extensive fibrosis and impaired hepatocyte regeneration. The only curative treatment is liver transplantation, but due to the several limitations of this procedure, the interest in alternative therapeutic strategies is increasing. In particular, the potential of bone marrow stem cell(BMSC) therapy in cirrhosis has been explored in different trials. In this article, we evaluate the results of 18 prospective clinical trials, and we provide a descriptive overview of recent advances in the research on hepatic regenerative medicine. The main message from the currently available data in the literature is that BMSC therapy is extremely promising in the context of liver cirrhosis. However, its application should be further explored in randomized, controlled trials with large cohorts and long follow-ups.

Clinical trial with traditional Chinese medicine intervention ''tonifying the kidney to promote liver regeneration and repair by affecting stem cells and their microenvironment'' for chronic hepatitis B-associated liver failure

AIM:To study the clinical efficacy of traditional Chinese medicine(TCM)intervention"tonifying the kidney to promote liver regeneration and repair by affecting stem cells and their microenvironment"("TTK")for treating liver failure due to chronic hepatitis B.METHODS:We designed the study as a randomized controlled clinical trial.Registration number of Chinese Clinical Trial Registry is Chi CTR-TRC-12002961.A total of 144 patients with liver failure due to infection with chronic hepatitis B virus were enrolled in this randomized controlled clinical study.Participants were randomly assigned to the following three groups:(1)a modern medicine control group(MMC group,36patients);(2)a"tonifying qi and detoxification"("TQD")group(72 patients);and(3)a"tonifying the kidney to promote liver regeneration and repair by affecting stem cells and their microenvironment"("TTK")group(36patients).Patients in the MMC group received general internal medicine treatment;patients in the"TQD"group were given a TCM formula"tonifying qi and detoxification"and general internal medicine treatment;patients in the"TTK"group were given a TCM formula of"TTK"and general internal medicine treatment.All participants were treated for 8 wk and then followed at 48 wk following their final treatment.The primaryefficacy end point was the patient fatality rate in each group.Measurements of various virological and biochemical indicators served as secondary endpoints.The one-way analysis of variance and the t-test were used to compare patient outcomes in the different treatment groups.RESULTS:At the 48-wk post-treatment time point,the patient fatality rates in the MMC,"TQD",and"TTK"groups were 51.61%,35.38%,and 16.67%,respectively,and the differences between groups were statistically significant(P<0.05).However,there were no significant differences in the levels of hepatitis B virus DNA or prothrombin activity among the three groups(P>0.05).Patients in the"TTK"group had significantly higher levels of serum total bilirubin compared to MMC subjects(339.40μmol/L±270.09μmol/L vs 176.13μmol/L±185.70μmol/L,P=0.014).Serum albumin levels were significantly increased in both the"TQD"group and"TTK"group as compared with the MMC group(31.30 g/L±4.77g/L,30.72 g/L±2.89 g/L vs 28.57 g/L±4.56 g/L,P<0.05).There were no significant differences in levels of alanine transaminase among the three groups(P>0.05).Safety data showed that there was one case of stomachache in the"TQD"group and one case of gastrointestinal side effect in the"TTK"group.CONCLUSION:Treatment with"TTK"improved the survival rates of patients with liver failure due to chronic hepatitis B.Additionally,liver tissue was regenerated and liver function was restored.

Therapeutic and regenerative potential of different sources of mesenchymal stem cells for cardiovascular diseases

Mesenchymalstemcells(MSCs)areidealcandidatesfortreatingmanycardiovasculardiseases.MSCscanmodify the internal cardiac microenvironment to facilitate their immunomodulatory and differentiation abilities,which are essential to restore heart function.MSCs can be easily isolated from different sources,including bone marrow,adipose tissues,umbilical cord,and dental pulp.MSCs from various sources differ in their regenerative and therapeutic abilities for cardiovascular disorders.In this review,we will summarize the therapeutic potential of each MSC source for heart diseases and highlight the possible molecular mechanisms of each source to restore cardiac function.

Targeted migration of mesenchymal stem cells modified with CXCR4 to acute failing liver improves liver regeneration

AIM: To improve the colonization rate of transplanted mesenchymal stem cells (MSCs) in the liver and effect of MSC transplantation for acute liver failure (ALF).

Influence of adriamycin on changes in Nanog, Oct-4, Sox2, ARID1 and Wnt5b expression in liver cancer stem cells

AIM: To determine the influence of Adriamycin (ADM) on the changes in Nanog, Oct4, Sox2, as well as, in ARID1 and Wnt5b expression in liver cancer stem cells.

Clinical translation of bioartificial liver support systems with human pluripotent stem cell-derived hepatic cells

There is currently a pressing need for alternative the-rapies to liver transplantation. The number of patients waiting for a liver transplant is substantially higher than the number of transplantable donor livers, resulting in a long waiting time and a high waiting list mortality. An extracorporeal liver support system is one possible approach to overcome this problem. However, the ideal cell source for developing bioartificial liver(BAL) support systems has yet to be determined. Recent advancements in stem cell technology allow researchers to generate highly functional hepatocyte-like cells from human pluripotent stem cells(h PSCs). In this mini-review, we summarize previous clinical trials with different BAL systems, and discuss advantages of and potential obstacles to utilizing h PSC-derived hepatic cells in clinical-scale BAL systems.

Bone marrow-derived mesenchymal stem cell therapy for decompensated liver cirrhosis:A meta-analysis

AIM: To assess the efficacy and safety of bone marrow-derived mesenchymal stem cell (BM-MSC) in the treatment of decompensated liver cirrhosis.

Effects of heme oxygenase-1-modified bone marrow mesenchymal stem cells on microcirculation and energy metabolism following liver transplantation

AIM To investigate the effects of heme oxygenase-1(HO-1)-modified bone marrow mesenchymal stem cells(BMMSCs)on the microcirculation and energy metabolism of hepatic sinusoids following reduced-size liver transplantation(RLT)in a rat model.METHODS BMMSCs were isolated and cultured in vitro using an adherent method,and then transduced with HO-1-bearing recombinant adenovirus to construct HO-1/BMMSCs.A rat acute rejection model following 50%RLT was established using a two-cuff technique.Recipients were divided into three groups based on the treatment received:normal saline(NS),BMMSCs and HO-1/BMMSCs.Liver function was examined at six time points.The levels of endothelin-1(ET-1),endothelial nitric-oxide synthase(e NOS),inducible nitric-oxide synthase(i NOS),nitric oxide(NO),and hyaluronic acid(HA)were detected using an enzyme-linked immunosorbent assay.The portal vein pressure(PVP)was detected by Power Lab ML880.The expressions of ET-1,i NOS,e NOS,and von Willebrand factor(v WF)protein in the transplanted liver were detected using immunohistochemistry and Western blotting.ATPase in the transplanted liver was detected by chemical colorimetry,and the ultrastructural changes were observed under a transmission electron microscope.RESULTS HO-1/BMMSCs could alleviate the pathological changes and rejection activity index of the transplanted liver,and improve the liver function of rats following 50%RLT,with statistically significant differences compared with those of the NS group and BMMSCs group(P<0.05).In term of the microcirculation of hepatic sinusoids:The PVP on POD7 decreased significantly in the HO-1/BMMSCs and BMMSCs groups compared with that of the NS group(P<0.01);HO-1/BMMSCs could inhibit the expressions of ET-1 and i NOS,increase the expressions of e NOS and inhibit amounts of NO production,and maintain the equilibrium of ET-1/NO(P<0.05);and HO-1/BMMSCs increased the expression of v WF in hepatic sinusoidal endothelial cells(SECs),and promoted the degradation of HA,compared with those of the NS group and BMMSCs group(P<0.05).In term of the energy metabolism of the transplanted liver,HO-1/BMMSCs repaired the damaged mitochondria,and improved the activity of mitochondrial aspartate aminotransferase(ASTm)and ATPase,compared with the other two groups(P<0.05).CONCLUSION HO-1/BMMSCs can improve the microcirculation of hepatic sinusoids significantly,and recover the energy metabolism of damaged hepatocytes in rats following RLT,thus protecting the transplanted liver.

Dynamic tracking of stem cells in an acute liver failure model

AIM： To investigate a dual labeling technique, which would enable real-time monitoring of transplanted em- bryonic stem cell （ESC） kinetics, as well as long-term tracking. METHODS： Liver damage was induced in C57/BL6 male mice （n = 40） by acetaminophen （APAP） 300 mg/kg administered intraperitoneally. Green fluores- cence protein （GFP） positive C57/BL6 mouse ESCs were stained with the near-infrared fluorescent lipophilic tracer 1,1-dioctadecyl-3,3,3,3-tetramethylindotricarbo- cyanine iodide （DiR） immediately before transplantationinto the spleen. Each of the animals in the cell therapy group （n = 20） received 5 x 106 ESCs 4 h following treatment with APAP. The control group （n = 20） re- ceived the vehicle only. The distribution and dynamics of the cells were monitored in real-time with the IVIS lumina-2 at 30 rain post transplantation, then at 3, 12, 24, 48 and 72 h, and after one and 2 wk. Immunohisto- chemical examination of liver tissue was used to identify expression of GFP and albumin. Plasma alanine amino- transferase （ALT） was measured as an indication of liver damage.RESULTS： DiR-stained ESCs were easily tracked with the IVIS using the indocyanine green filter due to its high background passband with minimal background autofluorescence. The transplanted cells were confined inside the spleen at 30 min post-transplantation, gradu- ally moved into the splenic vein, and were detectable in parts of the liver at the 3 h time-point. Within 24 h of transplantation, homing of almost 90% of cells was confirmed in the liver. On day three, however, the DiR signal started to fade out, and ex vivo IVIS imaging of different organs allowed signal detection at time-points when the signal could not be detected by in vivo imag- ing, and confirmed that the highest photon emission was in the liver （P 〈 0.0001）. At 2 wk, the DiRsignal was no longer detectable in vivo; however, immuno- histochemistry analysis of constitutively-expressed GFP was used to provide an insight into the distribution of the cells. GFP ＋ve cells were detected in tissue sections resembling hepatocytes and were dispersed throughout the hepatic parenchyma, with the presence of a larger number of GFP ＋ve cells incorporated within the sinu- soidal endothelial lining. Very faint albumin expression was detected in the transplanted GFP ＋re cells at 72 h; however at 2 wk, few cells that were positive for GFP were also strongly positive for albumin. There was a significant improvement in serum levels of ALT, albumin and bilirubin in both groups at 2 wk when compared with the 72 h time-point. In the cell therapy group, serum ALT was significantly （P = 0.016） lower and al- bumin （P = 0.009） was significantly higher when com- pared with the control group at the 2 wk time-point;however there was no difference in mortality between the two groups. CONCLUSION： Dual labeling is an easy to use and cheap method for longitudinal monitoring of distribu- tion, survival and engraftment of transplanted cells, and could be used for cell therapy models.