Hepatocyte Proliferation Research Articles

Ginkgetin exhibits antifibrotic effects by inducing hepatic stellate cell apoptosis via STAT1 activation.

Liver fibrosis affects approximately 800 million patients worldwide, with over 2 million deaths each year. Nevertheless, there are no approved medications for treating liver fibrosis. In this study, we investigated the impacts of ginkgetin on liver fibrosis and the underlying mechanisms. The impacts of ginkgetin on liver fibrosis were assessed in mouse models induced by thioacetamide or bile duct ligation. Experiments on human LX-2 cells and primary mouse hepatic stellate cells (HSCs) were performed to explore the underlying mechanisms, which were also validated in the mouse models. Ginkgetin significantly decreased hepatic extracellular matrix deposition and HSC activation in the fibrotic models induced by thioacetamide (TAA) and bile duct ligation (BDL). Beneficial effects also existed in inhibiting hepatic inflammation and improving liver function. In vitro experiments showed that ginkgetin markedly inhibited HSC viability and induced HSC apoptosis dose-dependently. Mechanistic studies revealed that the antifibrotic effects of ginkgetin depend on STAT1 activation, as the effects were abolished in vitro after STAT1 silencing and in vivo after inhibiting STAT1 activation by fludarabine. Moreover, we observed a meaningful cross-talk between HSCs and hepatocytes, in which IL-6, released by ginkgetin-induced apoptotic HSCs, enhanced hepatocyte proliferation by activating STAT3 signaling. Ginkgetin exhibits antifibrotic effects by inducing HSC apoptosis via STAT1 activation and enhances hepatocyte proliferation secondary to HSC apoptosis via the IL-6/STAT3 pathway.

Cow placenta extract ameliorates d-galactose-induced liver damage by regulating BAX/CASP3 and p53/p21/p16 pathways

Ethnopharmacological relevancePlacenta is a kind of traditional Chinese medicine, known as “Ziheche”, which has the function of tonifying qi and blood, nourishing liver and kidney. Placenta extract (PE) has been used for delaying organismal aging and treating various liver diseases. Cow placenta is a rich natural resource with large mass. Its composition is similar to that of human placenta, but it has not been effectively utilized. However, little is known about the effect of CPE on the liver of aging mice. Aim of the studyThe aim of this study is to explore the protective effect and mechanism of CPE on the liver of d-galactose (D-gal) induced aging mice. Materials and methodsStatistical methods were used to calculate mouse body weight and liver index. Hematoxylin-eosin (H&E) and transmission electron microscopy (TEM) were used to detect the morphological structure of the liver. Automatic biochemical analyzer was used to measure serum biochemical indicators. Three special staining methods were used to observe hepatocytes apoptosis, senescence and proliferation respectively. Relative kits were used to detect oxidative, inflammatory, and aging markers in the liver. Finally, real-time quantitative polymerase chain reaction and western-blot were used to detect aging related signaling pathways. ResultsCPE significantly improved the morphological damage and dysfunction of liver, restored the activities of liver enzymes in serum, and alleviated liver oxidative stress and inflammatory response in D-gal induced aging mice. Furthermore, CPE inhibited hepatocyte apoptosis and senescence, and promoted hepatocyte proliferation by regulating BAX/CASP3 and p53/p21/p16 signaling pathways, ultimately reduced the effects of aging on the liver. ConclusionCPE effectively ameliorated the impact of aging on the liver by inhibiting free radical production or scavenging excessive free radicals, and its mechanism is associated to the regulation of apoptosis and proliferation-related factors.

Acetaminophen overdose-induced acute liver injury can be alleviated by static magnetic field.

Acetaminophen (APAP), the most frequently used mild analgesic and antipyretic drug worldwide, is implicated in causing 46% of all acute liver failures in the USA and between 40% and 70% in Europe. The predominant pharmacological intervention approved for mitigating such overdose is the antioxidant N-acetylcysteine (NAC); however, its efficacy is limited in cases of advanced liver injury or when administered at a late stage. In the current study, we discovered that treatment with a moderate intensity static magnetic field (SMF) notably reduced the mortality rate in mice subjected to high-dose APAP from 40% to 0%, proving effective at both the initial liver injury stage and the subsequent recovery stage. During the early phase of liver injury, SMF markedly reduced APAP-induced oxidative stress, free radicals, and liver damage, resulting in a reduction in multiple oxidative stress markers and an increase in the antioxidant glutathione (GSH). During the later stage of liver recovery, application of vertically downward SMF increased DNA synthesis and hepatocyte proliferation. Moreover, the combination of NAC and SMF significantly mitigated liver damage induced by high-dose APAP and increased liver recovery, even 24 h post overdose, when the effectiveness of NAC alone substantially declines. Overall, this study provides a non-invasive non-pharmaceutical tool that offers dual benefits in the injury and repair stages following APAP overdose. Of note, this tool can work as an alternative to or in combination with NAC to prevent or minimize liver damage induced by APAP, and potentially other toxic overdoses.

A CDAHFD-induced mouse model mimicking human NASH in the metabolism of hepatic phosphatidylcholines and acyl carnitines.

Non-alcoholic steatohepatitis (NASH) is the hepatic manifestation of a cluster of conditions associated with lipid metabolism disorders. Ideal animal models mimicking the human NASH need to be explored to better understand the pathogenesis. A choline-deficient, L-amino acid-defined, high-fat diet (CDAHFD) has recently been used to induce the NASH model, but the advantages are not established. NASH models were induced using the well-known traditional methionine- and choline-deficient (MCD) diet for 5 weeks and the recently used CDAHFD for 3 weeks. Liver phenotypes were analyzed to evaluate the differences in markers related to NASH. Lipidomics and metabolism analyses were used to investigate the effects of dietary regimens on the lipidome of the liver. The CDAHFD induced stronger NASH responses than the MCD, including lipid deposition, liver injury, inflammation, bile acid overload and hepatocyte proliferation. A significant difference in the hepatic lipidome was revealed between the CDAHFD and MCD-induced NASH models. In particular, the CDAHFD reduced the hepatic levels of phosphatidylcholines (PCs) and acylcarnitines (ACs), which was supported by the metabolism analysis and in line with the tendency of human NASH. Pathologically, the CDAHFD could effectively induce a more human-like NASH model over the traditional MCD. The hepatic PCs, ACs and their metabolism in CDAHFD-treated mice were down-regulated, similar to those in human NASH.

Functions and Clinical Relevance of Liver-Derived Immunoglobulins.

Liver is the largest internal organ of the body with vital functions. In addition to its endocrine and exocrine activities, liver also plays a pivotal role in the immune system, including haematopoietic functions. Liver parenchymal cells, which are epithelial cells, have been found to possess innate immune functions by expressing pattern-recognition receptors (PRRs), producing complement components, and secreting cytokines. Intriguingly, in recent years, it has been discovered that liver epithelial cells also produce immunoglobulins (Igs), which have long been thought to be produced exclusively by B cells. Notably, even liver epithelial cells from B lymphocyte-deficient mice, including SCID mice and μMT mice, could also produce Igs. Compelling evidence has revealed both the physiological and pathological functions of liver-derived Igs. For instance, liver epithelial cells-derived IgM can serve as a source of natural and specific antibodies that contribute to innate immune responses, while liver-produced IgG can act as a growth factor to promote cell proliferation and survival in normal hepatocytes and hepatocarcinoma. Similar to that in B cells, the toll-like receptor 9 (TLR9)-MyD88 signaling pathway is also actively involved in promoting liver epithelial cells to secrete IgM. Liver-derived Igs could potentially serve as biomarkers, prognostic indicators, and therapeutic targets in the clinical setting, particularly for liver cancers and liver injury. Nevertheless, despite significant advances, much remains unknown about the mechanisms governing Ig transcription in liver cells, as well as the detailed functions of liver-derived Igs and their involvement in diseases and adaptive immunity. Further studies are still needed to reveal these underlying, undefined issues related to the role of liver-derived Igs in both immunity and diseases.

Clinical significance of CDT1 mRNA expression in chronic hepatitis C or liver cirrhosis.

We have previously reported that chromatin licensing and DNA replication factor 1 (CDT1) is associated with the postoperative recurrence of hepatocellular carcinoma (HCC). Based on this fact, we verified whether CDT1 mRNA expression is also associated with HCC development from chronic hepatitis C (CHC) and liver cirrhosis (LC). There were 142 cases with CHC or LC who underwent liver biopsy. Detection of CDT1 mRNA in liver was performed by RT-qPCR using frozen liver biopsy tissues. We examined the association between the CDT1 mRNA expression and clinical conditions and long-term outcome. We then examined the association between serum cytokine/chemokine levels and CDT1 mRNA expression in 58 cases. The cumulative incidence rates of HCC development in cases with CDT1 mRNA in the low expression group showed significantly lower than those in the high expression group (p = 0.0391). A significant correlation was found between CDT1 mRNA expression and the extent of proliferation of atypical hepatocytes in hematoxylin and eosin-stained sections (p<0.0001). CDT1 mRNA expression has been associated with cytokines involved in tumorigenesis in experimental and human cancers. We found that cases with high CDT1 mRNA expression were at risk for developing HCC, even if they were CHC or LC.

Intrahepatic neutrophil accumulation and extracellular trap formation are associated with posthepatectomy liver failure.

Posthepatectomy liver failure (PHLF) represents a life-threatening complication with limited therapeutic options. Neutrophils play a critical and dynamic role during regeneratory processes, but their role in human liver regeneration is incompletely understood, especially as underlying liver disease, detectable in the majority of patients, critically affects hepatic regeneration. Here we explored intrahepatic neutrophil accumulation and neutrophil extracellular traps (NETs) in patients with PHLF and validated the functional relevance of NETs in a murine partial hepatectomy (PHx) model. We investigated the influx of neutrophils, macrophages, eosinophils, and mast cells and the presence of their respective extracellular traps in liver biopsies of 35 patients undergoing hepatectomy (10 patients with PHLF) before and after the initiation of liver regeneration by fluorescence microscopy. In addition, NET formation and neutrophil activation were confirmed by plasma analysis of 99 patients (24 patients with PHLF) before and up to 5 days after surgery. Furthermore, we inhibited NETs via DNase I in a murine PHx model of mice with metabolically induced liver disease. We detected rapid intrahepatic neutrophil accumulation, elevated levels of myeloperoxidase release, and NET formation in regenerating human livers, with a significantly higher increase of infiltrating neutrophils and NETs in patients with PHLF. Circulating markers of neutrophil activation, including elastase, myeloperoxidase, and citrullinated histone H3, correlated with markers of liver injury. In a murine PHx model, we showed that the inhibition of NET accelerated hepatocyte proliferation and liver regeneration. Patients with PHLF showed accelerated intrahepatic neutrophil infiltration and NET formation, which were associated with liver damage. Further, we identified postsurgical myeloperoxidase levels as predictive markers for adverse outcomes and observed that blocking NETs in a murine PHx model accelerated tissue regeneration.

Activated Gab1 drives hepatocyte proliferation and anti-apoptosis in liver fibrosis via potential involvement of the HGF/c-Met signaling axis.

Chronic liver diseases are caused by hepatic viral infection, chemicals, and metabolic stress. The protein Grb2-associated binder 1 (Gab1) binds to various growth factor receptors, and triggers cell differentiation/survival signaling pathways. To identify signaling molecules involved in the progression of liver diseases, we performed reverse-phase protein microarray (RPMA)-based screening of hepatocytes isolated from humanized mice after acute HCV infection. Acute viral infection in humanized liver mice significantly decreased the level of hepatocyte p-Gab1. Moreover, hepatoma cells upon HCV infection decreased Gab1 mRNA at later times of infection (D3 to D5) and p-Gab1 level was inversely related to the production of TGF-β. In contrast, the level of p-Gab1 was increased in CCL4-induced fibrotic liver. Hepatoma cells showed elevation of p-Gab1, along with an increase in STAT3 and ERK activation, upon treatment with HGF (ligand of HGF receptor/c-Met) and CCL4. In Gab1 knockdown hepatoma cells, cell proliferative signaling activity was reduced but the level of activated caspase-3 was increased. These findings suggest that hepatocyte Gab1 expression may play a role in promoting liver fibrosis progression by triggering ERK activation and inhibiting apoptosis. It implies that the Gab1-mediated signaling pathway would be a promising therapeutic target to treat chronic liver diseases.

Analysis of Curcumin on proliferation, apoptosis and differentiation of hepatocyte mice balb/C exposure to benzapyrene: A Systematic Review

Objective: To analyse Curcumin effect on proliferation, apoptosis, and hepatocyte differentiation of Balb/C Mice exposed to benzapyrene. Methodology: A literature search was carried out systematically through the PubMed, NCBI, and Google Scholar databases using keywords, “Curcumin, Proliferation, Apoptosis and Hepatocyte Differentiation of Mice Balb/C, Benzapyrene”. Based on these keywords, the articles obtained were first selected by setting several inclusion criteria. Results: A total of 550 articles were obtained for the keywords “Curcumin, Proliferation, Apoptosis and Hepatocyte Differentiation of Mice Balb/C, Benzapyrene and 28 met the inclusion criteria. Curcumin reduced the hepatocyte proliferation index in experimental animals exposed to BaP and increased the hepatocyte apoptosis index in experimental animals exposed to BaP. It reduced the N/C ratio of hepatocytes in experimental animals exposed to BaP. Conclusion: Administration of curcumin in the diet during the initiation and post-initiation periods can inhibit the increase in PCNA expression, increase apoptosis, and improve hepatocyte differentiation patterns exposed to benzopyrene and curcumin.

Molecular mechanism of homologous lactic acid bacteria regulating liver cell injury of snakehead fish

In order to clarify the molecular mechanism of the homologous LAB regulating the liver cell injury of C. argus, this study used Fxr as the key target protein for the treatment of liver cell injury, used LCA to construct the liver cell injury model, and used the Fxr inhibitor GS to specifically block Fxr and its related signaling pathways will further explore the protective mechanism of homologous L. lactis L19 against liver injury in C. argus in vitro. The results showed that the LCA-induced liver cell injury model of C. argus was successfully constructed. The optimal concentration of LCA was 20 μM, and the optimal duration of action was 24 h. 1.0 × 108 CFU/mL L. lactis L19 ECP showed a protective effect on LCA-induced liver cell injury, remarkably decrease the levels of liver injury-related biochemical parameters AST, ALT, ALP and LDH (P < 0.05), and regulate the expressions of BA metabolism-related genes fxr, bsep, mrp2, shp, cyp7a1 and cyp8b1 and BA metabolism-related proteins Fxr, Bsep, Shp and Cyp7a1. It is revealed that L. lactis L19 is a possible molecular mechanism that regulates the liver cell injury of C. argus by activating Fxr and its related signal pathways. In vitro test results show that L. lactis L19 can enhance the proliferation of hepatocytes, promote the efflux and metabolism of toxic BA, inhibit the BA synthesis and uptake, and play a protective role in LCA-induced liver cell injury in C. argus. At the gene and protein levels, it was verified that L. lactis L19 can activate Fxr and its related signal pathways, and proved that the protective effects of L. lactis L19 on LCA-induced liver cell injury was blocked by GS. Thus, L. lactis L19, as a recognized probiotic, has a bright future in the treatment of liver injury-related diseases.

Efficient and reproducible generation of human induced pluripotent stem cell-derived expandable liver organoids for disease modeling

Genetic liver disease modeling is difficult because it is challenging to access patient tissue samples and to develop practical and relevant model systems. Previously, we developed novel proliferative and functional liver organoids from pluripotent stem cells; however, the protocol requires improvement for standardization and reproducible mass production. Here, we improved the method such that it is suitable for scalable expansion and relatively homogenous production, resulting in an efficient and reproducible process. Moreover, three medium components critical for long-term expansion were defined. Detailed transcriptome analysis revealed that fibroblast growth factor signaling, the essential pathway for hepatocyte proliferation during liver regeneration, was mainly enriched in proliferative liver organoids. Short hairpin RNA-mediated knockdown of FGFR4 impaired the generation and proliferation of organoids. Finally, glycogen storage disease type Ia (GSD1a) patient-specific liver organoids were efficiently and reproducibly generated using the new protocol. They well maintained disease-specific phenotypes such as higher lipid and glycogen accumulation in the liver organoids and lactate secretion into the medium consistent with the main pathologic characteristics of patients with GSD1a. Therefore, our newly established liver organoid platform can provide scalable and practical personalized disease models and help to find new therapies for incurable liver diseases including genetic liver diseases.

Influence of perfluoroorganic emulsion on morphometric parameters of the liver in a systemic inflammatory response (experimental study)

The objective was to study the effect of drug «Oxyphtem»1 on liver morphometric parameters in a systemic inflammatory response.Materials and methods. The experimental study was carried out on male Wistar rats (n = 26). The rats of the experimental group were injected with drug «Oxyphtem» intravenously once. Daily observation was carried out during14 days. Rats were taken out of the experiment on the 15th day under light ether anesthesia. Liver samples were fixed in 10 % neutral formalin in phosphate buffer for 24 hours. Histological preparations were studied using the Measure Pixels image analysis computer program based on a Leica 2000 light microscope.Results. In the intact group of rats, no violations of the liver parenchyma were detected: hepatocytes had clear boundaries, the lamellar structure was preserved and sinusoidal capillaries were not dilated. With the development of the systemic inflammatory response in the liver of rats of the control group, expansion and blood filling of the central veins and sinusoids, violation of the structure of the liver plates, and perinuclear edema of hepatocytes were noted. In the experimental group of rats with the use of drug «Oxyphtem», the preservation of the structure of the liver plates was observed, binuclear hepatocytes were found, and the sinusoids were not expanded. We hypothesize that hepatocyte proliferation and the increase in binuclear hepatic cells were indicative of the regenerative response to systemic inflammatory damage and metabolic demand.Conclusion. The use of drug «Oxyphtem» under conditions of the experimental systemic inflammatory response increases reparative regeneration and adaptation of the liver.

Constitutive androstane receptor agonist initiates metabolic activity required for hepatocite proliferation

Constitutive androstane receptor (CAR, NR1I3) activation by chemical compounds evokes liver hyperplasia in rodent. 1,4-Bis[2-(3,5-dichloropyridyloxy)] benzene (TCPOBOP), a mouse CAR agonist, is most commonly used to study chemically induced liver hyperplasia and hepatocyte proliferation in vivo. TCPOBOP is potent murine liver chemical mitogen, which induces rapid direct liver hyperplasia independent of liver injury. In recent years, a lot of data has been accumulated on the transcription program that characterizes TCPOBOP-induced hepatocyte proliferation. However, there are scarce data about metabolic requirements of hepatocytes dividing upon treatment with xenobiotics. In present study, we employed liquid chromatography - mass spectrometry technology combined with statistical analysis to develop a metabolite profile of small biomolecules, to identify key metabolic changes in male mouse liver tissue after TCPOBOP administration. Analysis of biochemical pathways of the differentially affected metabolites in mouse livers demonstrated significant TCPOBOP-mediated enrichment of several processes including those relevant to nucleotide metabolism, amino acid metabolism, and energy substrate metabolism. Our findings provide evidence to support the conclusion that CAR agonist, TCPOBOP, initiates an intracellular program that promotes the global coordinated metabolic activities required for hepatocyte proliferation. Our metabolic data may provide novel insight into the biological mechanisms that occur during TCPOBOP-induced hepatocyte proliferation in mice.

In vivo genotoxicity testing strategies: Report from the 8th International Workshop on Genotoxicity Testing (IWGT)

AbstractThe in vivo working group (WG) considered three topics: acceptable maximum doses for negative erythrocyte micronucleus (MN) tests, validation status of MN assays in non‐hematopoietic tissues, and nuisance factors in the comet assay. The WG reached agreement on many issues, including: negative erythrocyte MN studies should be acceptable if dosing is conducted to Organisation for Economic Co‐operation and Development (OECD) test guideline (TG) 474 recommendations and if sufficient bone marrow exposure is demonstrated; consensus on the evidence required to demonstrate “sufficient” exposure was not reached. The liver MN test using six‐week‐old rats is sufficiently validated to develop an OECD TG, but the impact of animal age warrants additional study. Ki‐67 is a reliable marker for cellular proliferation in hepatocytes. The gastrointestinal tract MN test is useful for detecting poorly absorbed or rapidly degraded aneugens, and for genotoxic metabolites formed in the colon. Although current validation data are insufficient to support the development of an OECD TG, the methodologies are sufficient to consider as an appendix to OECD TG474. Comparison of comet assay results to laboratory historical control data (HCD) should not be used in data evaluation, unless the HCD distribution is demonstrated to be stable and the predominant source of HCD variation is due to animal, not study, factors. No universally acceptable negative control limit for any tissue was identified. Methodological differences in comet studies can result in variable data interpretations; more data are required before best practice recommendations can be made. Hedgehogs alone are unreliable indicators of cytotoxicity and additional investigations into cytotoxicity markers are required.

Fully umbilical cord-derived adhesive materials enable to recruit and segregate immune cells for the reversal of acute liver failure

Inflammatory cytokine storms can trigger disease exacerbation and even death and have reached a consensus in the clinical treatment of acute organ failure. However, the existing strategies remain a great challenge to efficiently suppress inflammatory cytokine storms for promoting organ repair and regeneration. Herein, fully human umbilical cord (UC)-derived adhesive materials (UCAM) that integrate decellularized extracellular matrix (ECM) nanofiber hydrogel and homologous mesenchymal stem cells (MSCs) are demonstrated to greatly suppress inflammatory cytokine storms, demonstrating high efficacy in treating acute liver failure (ALF) in rats with 90% hepatectomy. The UC-derived adhesive materials have the capacity to secrete a significant quantity of cytokines by MSCs to recruit activated immune cells to migrate into their ECM nanofiber networks, segregating them away from the infection area and thereby greatly suppressing the inflammatory cytokine storms. As expected, the UC-derived adhesive materials can significantly promote hepatocyte proliferation to achieve functional recovery and regeneration of the liver, significantly improving the survival rate in rats. Our fully human UC-derived adhesive materials provide a new avenue in suppressing inflammatory cytokine storms for promoting organ regeneration that would be really utility in clinical organ transplantation-related treatment.

Bifidobacterium longum promotes postoperative liver function recovery in patients with hepatocellular carcinoma

Timely liver function recovery (LFR) is crucial for postoperative hepatocellular carcinoma (HCC) patients. Here, we established the significance of LFR on patient long-term survival through retrospective and prospective cohorts and identified a key gut microbe, Bifidobacterium longum, depleted in patients with delayed recovery. Fecal microbiota transfer from HCC patients with delayed recovery to mice similarly impacted recovery time post hepatectomy. However, oral gavage of B.longum improved liver function and repair in these mice. In a clinical trial of HCC patients, orally administering a probiotic bacteria cocktail containing B.longum reduced the rates of delayed recovery, shortened hospital stays, and improved overall 1-year survival. These benefits, attributed to diminished liver inflammation, reduced liver fibrosis, and hepatocyte proliferation, were associated with changes in key metabolic pathways, including 5-hydroxytryptamine, secondary bile acids, and short-chain fatty acids. Our findings propose that gut microbiota modulation can enhance LFR, thereby improving postoperative outcomes for HCC patients.

Disrupting Poly(ADP-ribosyl)ating Pathway Creates Premalignant Conditions in Mammalian Liver.

Hepatocellular carcinoma (HCC) is a major global health concern, representing one of the leading causes of cancer-related deaths. Despite various treatment options, the prognosis for HCC patients remains poor, emphasizing the need for a deeper understanding of the factors contributing to HCC development. This study investigates the role of poly(ADP-ribosyl)ation in hepatocyte maturation and its impact on hepatobiliary carcinogenesis. A conditional Parg knockout mouse model was employed, utilizing Cre recombinase under the albumin promoter to target Parg depletion specifically in hepatocytes. The disruption of the poly(ADP-ribosyl)ating pathway in hepatocytes affects the early postnatal liver development. The inability of hepatocytes to finish the late maturation step that occurs early after birth causes intensive apoptosis and acute inflammation, resulting in hypertrophic liver tissue with enlarged hepatocytes. Regeneration nodes with proliferative hepatocytes eventually replace the liver tissue and successfully fulfill the liver function. However, early developmental changes predispose these types of liver to develop pathologies, including with a malignant nature, later in life. In a chemically induced liver cancer model, Parg-depleted livers displayed a higher tendency for hepatocellular carcinoma development. This study underscores the critical role of the poly(ADP-ribosyl)ating pathway in hepatocyte maturation and highlights its involvement in liver pathologies and hepatobiliary carcinogenesis. Understanding these processes may provide valuable insights into liver biology and liver-related diseases, including cancer.

Periportal hepatocyte proliferation at midgestation governs maternal glucose homeostasis in mice

The maternal liver is challenged by metabolic demands throughout pregnancy. However, hepatocyte dynamics and their physiological significance in pregnancy remain unclear. Here, we show in mice that hepatocyte proliferation is spatiotemporally regulated in each liver lobular zone during pregnancy, with transient proliferation of periportal and pericentral hepatocytes during mid and late gestation, respectively. Using adeno-associated virus (AAV)−8-mediated expression of the cell cycle inhibitor p21 in hepatocytes, we show that inhibition of hepatocyte proliferation during mid, but not late, gestation impairs liver growth. Transcriptionally, genes involved in glucose/glycogen metabolism are downregulated in late pregnancy when midgestational hepatocyte proliferation is attenuated. In addition, hepatic glycogen storage is abolished, with concomitant elevated blood glucose concentrations, glucose intolerance, placental glycogen deposition, and fetal overgrowth. Laser capture microdissection and RNA-seq analysis of each liver lobular zone show zone-specific changes in the transcriptome during pregnancy and identify genes that are periportally expressed at midgestation, including the hyaluronan-mediated motility receptor (Hmmr). Knockdown of Hmmr in hepatocytes by AAV8-shHmmr suppresses periportal hepatocyte proliferation at midgestation and induces impaired hepatic glycogen storage, glucose intolerance, placental glycogen deposition and fetal overgrowth. Our results suggest that periportal hepatocyte proliferation during midgestation is critical for maternal glycogen metabolism and fetal size.

Liver histology and hepatic progenitor cell activity in pediatric acute liver failure: Implications for clinical outcome.

Hepatic progenitor cell (HPC) activity and regenerative process that follows pediatric acute liver failure (PALF) is still not well understood. This clinicopathological study was thus conducted with an aim to study the correlation of liver histology and HPC activity with outcomes in PALF. All PALF patients with available hepatic histological specimens were included and specimens were analyzed for hepatocyte loss, HPC activity [using cytokeratin (CK) 7, CK19, sex-determining region Y-related high mobility group box(SOX)9 and epithelial cell adhesion molecule (EpCAM)], hepatocyte proliferation (using Ki67), and hepatocyte senescence (using p53 and p21). Ninety-four children were included: 22 (23.4%) survived with native liver (SNL) (i.e., the good outcome group) while rest (i.e., the poor outcome group) either died [33%, 35.1%] or received liver transplant (LT) [39%, 41.5%]. When compared to subjects with poor outcomes, those in the SNL group exhibited significantly less severe hepatocyte loss, fewer HPC/hpf, more proliferating hepatocytes, and less senescent hepatocytes (p < .05). Increasing severity of hepatocyte loss (adjusted OR: 9.95, 95% CI: 4.22-23.45, p < .001) was identified as an independent predictor of poor outcome. Eighty percent children with >50% native hepatocyte loss had poor outcome within 10 days of hospitalization. In PALF, more severe hepatocyte loss, higher number of HPC activation, lesser number of proliferating hepatocytes, and greater number of senescent hepatocytes are associated with a poor outcome. Loss of >50% hepatocytes is an independent predictor of poor outcome in PALF.

VEGFA mRNA-LNP promotes biliary epithelial cell-to-hepatocyte conversion in acute and chronic liver diseases and reverses steatosis and fibrosis

The liver is known for its remarkable regenerative ability through proliferation of hepatocytes. Yet, during chronic injury or severe hepatocyte death, proliferation of hepatocytes is exhausted. To overcome this hurdle, we propose vascular-endothelial-growth-factor A (VEGFA) as a therapeutic means to accelerate biliary epithelial-cell (BEC)-to-hepatocyte conversion. Investigation in zebrafish establishes that blocking VEGF receptors abrogates BEC-driven liver repair, while VEGFA overexpression promotes it. Delivery of VEGFA via nonintegrative and safe nucleoside-modified mRNA encapsulated into lipid nanoparticles (mRNA-LNPs) in acutely or chronically injured mouse livers induces robust BEC-to-hepatocyte conversion and elimination of steatosis and fibrosis. In human and murine diseased livers, we further identified VEGFA-receptor KDR-expressing BECs associated with KDR-expressing cell-derived hepatocytes. This work defines KDR-expressing cells, most likely being BECs, as facultative progenitors. This study reveals unexpected therapeutic benefits of VEGFA delivered via nucleoside-modified mRNA-LNP, whose safety is widely validated with COVID-19 vaccines, for harnessing BEC-driven repair to potentially treat liver diseases.