Dysregulation of cofilin-1 activity-the missing link between herpes simplex virus type-1 infection and Alzheimer's disease.

Alzheimer's disease (AD) is a multifactorial disease triggered by environmental factors in combination with genetic predisposition. Infectious agents, in particular herpes simplex virus type 1 (HSV-1), are gradually being recognised as important factors affecting the development of AD. However, the mechanism linking HSV-1 and AD remains unknown. Of note, HSV-1 manipulates the activity of cofilin-1 to ensure their efficient infection in neuron cells. Cofilin-1, the main regulator of actin cytoskeleton reorganization, is implicating for the plastic of dendritic spines and axon regeneration of neuronal cells. Moreover, dysfunction of cofilin-1 is observed in most AD patients, as well as in mice with AD and ageing. Further, inhibition of cofilin-1 activity ameliorates the host cognitive impairment in an animal model of AD. Together, dysregulation of cofilin-1 led by HSV-1 infection is a potential link between HSV-1 and AD. Herein, we critically summarize the role of cofilin-1-mediated actin dynamics in both HSV-1 infection and AD, respectively. We also propose several hypotheses regarding the connecting roles of cofilin-1 dysregulation in HSV-1 infection and AD. Our review provides a foundation for future studies targeting individuals carrying HSV-1 in combination with cofilin-1 to promote a more individualised approach for treatment and prevention of AD.

Anti-HSV-1 activity of Aspergillipeptide D, a cyclic pentapepetide isolated from fungus Aspergillus sp. SCSIO 41501.

BACKGROUND: Herpes simplex virus 1, an enveloped DNA virus belonging to the Herpesviridae family, spreads to neurons and causes pathological changes in the central nervous system. The purpose of this study was to investigate the potency and mechanism of antiviral activity of Aspergillipeptide D, a cyclic pentapeptide isolated from a culture broth of marine gorgonian-derived fungus Aspergillus sp. SCSIO 41501, At present, there are many studies on the anti-tumor, anti-clotting, anti-oxidant and immunoinflammatory effects of Aspergillipeptide D, but little research has been done on the anti-HSV-1 activity of Aspergillipeptide D. METHODS: The anti-HSV-1 activity of Aspergillipeptide D was evaluated by plaque reduction assay. The mechanism of action against HSV-1 was determined from the effective stage. Then we assayed the viral DNA replication, viral RNA synthesis and protein expression, respectively. We also identified the proteins that interact with gB by mass spectrometry, and assayed the effect of Aspergillipeptide D on the interaction between the virus gB protein and cell proteins. RESULTS: Plaque reduction experiments showed that Aspergillipeptide D did not affect HSV-1 early infection events, including viral inactivation, attachment and penetration. Interestingly, Aspergillipeptide D dramatically reduced both the gene and protein levels of viral late protein gB, and suppressed its location in the endoplasmic reticulum and Golgi apparatus. In contrast, overexpression of gB restored viral production. Finally, proteomic analysis revealed that the numbers of cellular proteins that interacted with gB protein was largely decreased by Aspergillipeptide D. These results suggested that Aspergillipeptide D inhibited gB function to affect HSV-1 intercellular spread. CONCLUSIONS: Our results indicated that Aspergillipeptide D might be a potential candidate for HSV-1 therapy, especially for ACV-resistant strains.

Correction to: Anti-HSV-1 activity of Aspergillipeptide D, a cyclic pentapeptide isolated from fungus Aspergillus sp. SCSIO 41501.

Following publication of the original article [1], we have been notified that there is a typo in the title of this article.

A comprehensive investigation of the mRNA and protein level of ACE2, the putative receptor of SARS-CoV-2, in human tissues and blood cells.

The outbreak of pneumonia caused by SARS-CoV-2 posed a great threat to global human health, which urgently requires us to understand comprehensively the mechanism of SARS-CoV-2 infection. Angiotensin-converting enzyme 2 (ACE2) was identified as a functional receptor for SARS-CoV-2, distribution of which may indicate the risk of different human organs vulnerable to SARS-CoV-2 infection. Previous studies investigating the distribution of ACE2 mRNA in human tissues only involved a limited size of the samples and a lack of determination for ACE2 protein. Given the heterogeneity among humans, the datasets covering more tissues with a larger size of samples should be analyzed. Indeed, ACE2 is a membrane and secreted protein, while the expression of ACE2 in blood and common blood cells remains unknown. Herein, the proteomic data in HIPED and the antibody-based immunochemistry result in HPA were collected to analyze the distribution of ACE2 protein in human tissues. The bulk RNA-seq profiles from three separate public datasets including HPA tissue Atlas, GTEx, and FANTOM5 CAGE were also obtained to determine the expression of ACE2 in human tissues. Moreover, the abundance of ACE2 in human blood and blood cells was determined by analyzing the data in the PeptideAtlas and the HPA Blood Atlas. We found that the mRNA expression cannot reflect the abundance of ACE2 factor due to the strong differences between mRNA and protein quantities of ACE2 within and across tissues. Our results suggested that ACE2 protein is mainly expressed in the small intestine, kidney, gallbladder, and testis, while the abundance of which in brain-associated tissues and blood common cells is low. HIPED revealed enrichment of ACE2 protein in the placenta and ovary despite a low mRNA level. Further, human secretome shows that the average concentration of ACE2 protein in the plasma of males is higher than those in females. Our research will be beneficial for understanding the transmission routes and sex-based differences in susceptibility of SARS-CoV-2 infection.

Roles of HSV-1 infection-induced microglial immune responses in CNS diseases: friends or foes?

Microglia, as brain-resident macrophages, are the first line of defense against brain invading pathogens. Further, their dysfunction has been recognized to be closely associated with mounting CNS diseases. Of note, chronic HSV-1 infection leads to the persistent activation of microglia, which elicit a comprehensive response by generating certain factors with neurotoxic and neuroprotective effects. CNS infection with HSV-1 results in herpes simplex encephalitis and herpes simplex keratitis. Microglial immune response plays a crucial role in the development of these diseases. Moreover, HSV-1 infection is strongly associated with several CNS diseases, especially Alzheimer's disease and schizophrenia. These CNS diseases can be effectively ameliorated by eliciting an appropriate immune response, such as inhibition of microglial proliferation and activation. Therefore, it is crucial to reassess the positive and negative roles of microglia in HSV-1 CNS infection for a more comprehensive and detailed understanding of the relationship between microglia and CNS diseases. Hence, the present review focuses on the dual roles of microglia in mediating HSV-1 CNS infection, as well as on the strategy of targeting microglia to ameliorate CNS diseases. Further research in this field can help comprehensively elucidate the dual role of the microglial immune response in HSV-1 CNS infection, providing a theoretical basis for identifying therapeutic targets against overactive microglia in CNS diseases and HSV-1 infection.

Microfluidic Synthesis of CuH Nanoparticles for Antitumor Therapy through Hydrogen-Enhanced Apoptosis and Cuproptosis.

Cuproptosis has drawn enormous attention in antitumor material fields; however, the responsive activation of cuproptosis against tumors using nanomaterials with high atom utilization is still challenging. Herein, a copper-based nanoplatform consisting of acid-degradable copper hydride (CuH) nanoparticles was developed via a microfluidic synthesis. After coating with tumor-targeting hyaluronic acid (HA), the nanoplatform denoted as HA-CuH-PVP (HCP) shows conspicuous damage toward tumor cells by generating Cu+ and hydrogen (H2) simultaneously. Cu+ can induce apoptosis by relying on Fenton-like reactions and lead to cuproptosis by causing mitochondrial protein aggregation. Besides, the existence of H2 can enhance both cell death types by causing mitochondrial dysfunction and intracellular redox homeostatic disorders. In vivo experimental results further exhibit the desirable potential of HCP for killing tumor cells and inhibiting lung metastases, which will broaden the horizons of designing copper-based materials triggering apoptosis and cuproptosis for better antitumor efficacy.

Self-Generating Gold Nanocatalysts in Autologous Tumor Cells for Targeted Catalytic Immunotherapy.

Employing tumor whole cells for tumor immunotherapy is a promising tumor therapy proposed in the early stage, but its therapeutic efficacy is weakened by the methods of eliminating pathogenicity and the mass ratio of the effective antigen carried by itself. Here, by adding gold ion to live cancer cells in the microfluidic droplets, this work obtains dead tumor whole cells with NIR-controlled catalytic ability whose pathogenicity is removed while plenary tumor antigens, major structure, and homing ability are reserved. The engineered tumor cell (Cell-Au) with the addition of prodrug provides 1O2 in an O2-free Russell mechanism, which serves better in a hypoxic tumor microenvironment. This tumor whole-cell catalytic vaccine (TWCV) promotes the activation of dendritic cells and the transformation of macrophages into tumor suppressor phenotype. In 4T1 tumor-bearing mice, the Cell-Au-based vaccine supports the polarization of cytotoxicity T cells, resulting in tumor eradication and long-term animal survival. Compared with antigen vaccines or adoptive cell therapy which takes months to obtain, this TWCV can be prepared in just a few days with satisfactory immune activation and tumor therapeutic efficacy, which provides an alternative way for the preparation of personalized tumor vaccines across tumor types and gives immunotherapy a new path.

Logic-gated tumor-microenvironment nanoamplifier enables targeted delivery of CRISPR/Cas9 for multimodal cancer therapy.

Recent innovations in nanomaterials inspire abundant novel tumor-targeting CRISPR-based gene therapies. However, the therapeutic efficiency of traditional targeted nanotherapeutic strategies is limited by that the biomarkers vary in a spatiotemporal-dependent manner with tumor progression. Here, we propose a self-amplifying logic-gated gene editing strategy for gene/H2O2-mediated/starvation multimodal cancer therapy. In this approach, a hypoxia-degradable covalent-organic framework (COF) is synthesized to coat a-ZIF-8 in which glucose oxidase (GOx) and CRISPR system are packaged. To intensify intracellular redox dyshomeostasis, DNAzymes which can cleave catalase mRNA are loaded as well. When the nanosystem gets into the tumor, the weakly acidic and hypoxic microenvironment degrades the ZIF-8@COF to activate GOx, which amplifies intracellular H+ and hypoxia, accelerating the nanocarrier degradation to guarantee available CRISPR plasmid and GOx release in target cells. These tandem reactions deplete glucose and oxygen, leading to logic-gated-triggered gene editing as well as synergistic gene/H2O2-mediated/starvation therapy. Overall, this approach highlights the biocomputing-based CRISPR delivery and underscores the great potential of precise cancer therapy.

Self-Metallized Whole Cell Vaccines Prepared by Microfluidics for Bioorthogonally Catalyzed Antitumor Immunotherapy.

Tumor whole cell, carrying a complete set of tumor-associated antigens and tumor-specific antigens, has shown great potential in the construction of tumor vaccines but is hindered by the complex engineering means and limited efficacy to cause immunity. Herein, we provided a strategy for the self-mineralization of autologous tumor cells with palladium ions in microfluidic droplets, which endowed the engineered cells with both immune and catalytic functions, to establish a bioorthogonally catalytic tumor whole-cell vaccine. This vaccine showed strong inhibition both in the occurrence and recurrence of tumor by invoking the immediate antitumor immunity and building a long-term immunity.

Oleanolic Acid Derivative AXX-18 Exerts Antiviral Activity by Inhibiting the Expression of HSV-1 Viral Genes UL8 and UL52.

Two-thirds of the world's population is infected with HSV-1, which is closely associated with many diseases, such as Gingival stomatitis and viral encephalitis. However, the drugs that are currently clinically effective in treating HSV-1 are Acyclovir (ACV), Ganciclovir, and Valacyclovir. Due to the widespread use of ACV, the number of drug-resistant strains of ACV is increasing, so searching for new anti-HSV-1 drugs is urgent. The oleanolic-acid derivative AXX-18 showed a CC50 value of 44.69 µM for toxicity to HaCaT cells and an EC50 value of 1.47 µM for anti-HSV-1/F. In addition, AXX-18 showed significant inhibition of ACV-resistant strains 153, 106, and Blue, and the anti-HSV-1 activity of AXX-18 was higher than that of oleanolic acid. The mechanism of action of AXX-18 was found to be similar to that of oleanolic acid, except that AXX-18 could act on both the UL8 and UL52 proteins of the uncoupling helicase-primase enzyme, whereas oleanolic acid could only act on the UL8 protein. We have elucidated the antiviral mechanism of AXX-18 in detail and, finally, found that AXX-18 significantly inhibited the formation of skin herpes. In conclusion, we have explored the anti-HSV-1 activity of AXX-18 in vitro and in vivo as well as identification of its potential target proteins, which will provide a theoretical basis for the development of subsequent anti-HSV-1 drugs.

Herpes Simplex Virus 1 Induces Microglia Gasdermin D-Dependent Pyroptosis Through Activating the NLR Family Pyrin Domain Containing 3 Inflammasome.

Herpes simplex virus type 1 (HSV-1) is a highly prevalent virus in humans and causes severe forms of inflammation, such as herpes simplex encephalitis (HSE). Pyroptosis is a new inflammatory cell death triggered by inflammasome and cysteine-requiring aspartate protease-1 (caspase-1) activation. Nonetheless, HSV-1 induces encephalitis, and cell death mechanisms are not understood. In this study, we confirmed for the first time that the DNA virus HSV-1 triggers Gasdermin D-dependent pyroptosis by activating NLR family pyrin domain containing 3 (NLRP3) inflammasomes in mouse microglia, leading to mature IL-1ß production and active caspase-1 (p10) release. Inhibition of microglial NLRP3 inflammasome activation suppressed HSV-1-induced Gasdermin D-dependent pyroptosis. In addition, NLRP3 and IL-1ß expression levels were significantly increased in the mouse model of herpes simplex encephalitis compared with normal mice without viral infection. Collectively, our data revealed that the activation of inflammasomes and GSDMD-dependent pyroptosis is the mechanism of HSV-1 inducing inflammation and provides treatment targets for viral inflammation.

Hsp90 Inhibitors Prevent HSV-1 Replication by Directly Targeting UL42-Hsp90 Complex.

Herpes simplex virus type I (HSV-1) is a member of the Alphaherpesvirinae family, which could initiate labial herpes caused by the reactivation of HSV-1 primary infection, and secondary infection even causes herpes encephalitis. The study presented here demonstrates that Hsp90 inhibitors (AT-533 and 17-AAG) directly targeted the HSV-1 UL42-Hsp90 complex, and Hsp90 interacted with HSV-1 UL42 in silicon and experiment. Interestingly, Hsp90 inhibitors also reduced virus titers of ACV-resistant clinical HSV-1 strains (153 and blue strain), revealing that HSV-1 UL42 would be a new target against ACV-resistant HSV-1 strains. Altogether, this present study indicates that Hsp90 inhibitors prevent HSV-1 proliferation by regulating the interaction between Hsp90 and HSV-1 UL42, suggesting a promising target for anti-HSV-1 therapies in the replication.

Hsp90 Inhibitors Inhibit the Entry of Herpes Simplex Virus 1 Into Neuron Cells by Regulating Cofilin-Mediated F-Actin Reorganization.

Herpes simplex virus 1 (HSV-1) is a common neurotropic virus, the herpes simplex encephalitis (HSE) caused by which is considered to be the most common sporadic but fatal encephalitis. Traditional antiviral drugs against HSV-1 are limited to nucleoside analogs targeting viral factors. Inhibition of heat shock protein 90 (Hsp90) has potent anti-HSV-1 activities via numerous mechanisms, but the effects of Hsp90 inhibitors on HSV-1 infection in neuronal cells, especially in the phase of virus entry, are still unknown. In this study, we aimed to investigate the effects of the Hsp90 inhibitors on HSV-1 infection of neuronal cells. Interestingly, we found that Hsp90 inhibitors promoted viral adsorption but inhibited subsequent penetration in neuronal cell lines and primary neurons, which jointly confers the antiviral activity of the Hsp90 inhibitors. Mechanically, Hsp90 inhibitors mainly impaired the interaction between Hsp90 and cofilin, resulting in reduced cofilin membrane distribution, which led to F-actin polymerization to promote viral attachment. However, excessive polymerization of F-actin inhibited subsequent viral penetration. Consequently, unidirectional F-actin polymerization limits the entry of HSV-1 virions into neuron cells. Our research extended the molecular mechanism of Hsp90 in HSV-1 infection in neuron cells and provided a theoretical basis for developing antiviral drugs targeting Hsp90.

Subacute toxicological evaluation of AT-533 and AT-533 gel in Sprague-Dawley rats.

As a novel heat shock protein 90 inhibitor, AT-533 exhibits various biological activities in vitro, including anti-viral, anti-tumor and anti-inflammatory activities. Moreover, AT-533 gel, a gel dosage form of AT-533, has been suggested to have anti-keratitis and herpes simplex virus type-1 infection-induced effects on the skin lesions of animals. However, the safety evaluation of AT-533 and AT-533 gel has, to the best of our knowledge, not been examined in in vivo toxicological tests. Therefore, these toxicological tests were carried out in the present study. A 30-day subacute toxicity test for AT-533 was conducted at doses of 1, 2 and 4 mg/kg in Sprague-Dawley rats, while that for AT-533 gel was conducted using a single dose of 5 g/kg. The toxicological tests showed that a high-dose of AT-533 caused lethality and side effects in Sprague-Dawley rats. However, no mortality, loss of appetite and body weight, adverse reactions, or toxicologically relevant alterations in hematology, biochemistry and macroscopic findings (except for skin) occurred in rats exposed to low-dose AT-533 and single-dose AT-533 gel (5 g/kg) during a 30-day subacute dermic toxicity study. The aforementioned results suggested that AT-533 gel is non-toxic for Sprague-Dawley rats, as shown by a dermic subacute toxicity test and that except for slight skin irritation, AT-533 gel had almost no side effects when administered percutaneously for 30 days.

Single-cell RNA-sequencing analysis identifies host long noncoding RNA MAMDC2-AS1 as a co-factor for HSV-1 nuclear transport.

Herpes simplex virus (HSV) type 1 (HSV-1) infection exhibited high heterogeneity at individual cells level, including the different gene expression patterns and varying amounts of progeny virus. However, the underlying mechanism of such variability remains obscure. The importance of host long noncoding RNAs (lncRNAs) in virus infection had been recognized, while the contribution of lncRNAs to the heterogeneous infection remains unknown. Herein, a prior single-cell RNA sequencing data using HSV-1 reporter strain expressing ICP4-YFP was re-analyzed to obtain the differentially expressed lncRNA between the successfully initiated viral gene expression (ICP4-YFP+) cells and the aborted infection cells (ICP4-YFP-). The ICP4-YFP+ population show a higher abundance of MAMDC2 antisense 1 (MAMDC2-AS1) lncRNA than ICP4-YFP- population. MAMDC2-AS1 silencing reduces the expression of HSV-1 immediate early (IE) genes and limit HSV-1 infection in human host cells. Consistently, ectopic expression of MAMDC2-AS1 enhances HSV-1 IE genes transcription and facilitates the formation of HSV-1-induced plaques. Mechanically, both RNA-pull down and RNA immunoprecipitation assays show that MAMDC2-AS1 interacts with the RNA binding protein heat shock protein 90α (Hsp90α), a molecular chaperone involving in the nuclear import of HSV-1. The MAMDC2-AS1-Hsp90α interaction facilitates the nuclear transport of viral tegument protein VP16, the core factor initiating the expression of HSV-1 IE genes. The transcription factor YY1 mediates the induction of MAMDC2-AS1 upon HSV-1 infection. Our study elucidates the contribution of lncRNA to HSV-1 infection susceptibility in human cells and the role of Hsp90α RNA binding activity in HSV-1 infection.

Intestinal Microbiota-Associated Metabolites: Crucial Factors in the Effectiveness of Herbal Medicines and Diet Therapies.

Although the efficacy of herbal medicines (HMs) and traditional Chinese medicines (TCMs) in human diseases has long been recognized, their development has been hindered in part by a lack of a comprehensive understanding of their mechanisms of action. Indeed, most of the compounds extracted from HMs can be metabolized into specific molecules by host microbiota and affect pharmacokinetics and toxicity. Moreover, HMs modulate the constitution of host intestinal microbiota to maintain a healthy gut ecology. Dietary interventions also show great efficacy in treating some refractory diseases, and the commensal microbiota potentially has significant implications for the high inter-individual differences observed in such responses. Herein, we mainly discuss the contribution of the intestinal microbiota to high inter-individual differences in response to HMs and TCMs, and especially the already known metabolites of the HMs produced by the intestinal microbiota. The contribution of commensal microbiota to the inter-individual differences in response to dietary therapy is also briefly discussed. This review highlights the significance of intestinal microbiota-associated metabolites to the efficiency of HMs and dietary interventions. Our review may help further identify the mechanisms leading to the inter-individual differences in the effectiveness of HM and dietary intervention from the perspective of their interactions with the intestinal microbiota.

The Gut-Microglia Connection: Implications for Central Nervous System Diseases.

The importance of the gut microbiome in central nervous system (CNS) diseases has long been recognized; however, research into this connection is limited, in part, owing to a lack of convincing mechanisms because the brain is a distant target of the gut. Previous studies on the brain revealed that most of the CNS diseases affected by the gut microbiome are closely associated with microglial dysfunction. Microglia, the major CNS-resident macrophages, are crucial for the immune response of the CNS against infection and injury, as well as for brain development and function. However, the current understanding of the mechanisms controlling the maturation and function of microglia is obscure, especially regarding the extrinsic factors affecting microglial function during the developmental process. The gut microflora has been shown to significantly influence microglia from before birth until adulthood, and the metabolites generated by the microbiota regulate the inflammation response mediated by microglia in the CNS; this inspired our hypothesis that microglia act as a critical mediator between the gut microbiome and CNS diseases. Herein, we highlight and discuss current findings that show the influence of host microbiome, as a crucial extrinsic factor, on microglia within the CNS. In addition, we summarize the CNS diseases associated with both the host microbiome and microglia and explore the potential pathways by which the gut bacteria influence the pathogenesis of CNS diseases. Our work is thus a comprehensive theoretical foundation for studies on the gut-microglia connection in the development of CNS diseases; and provides great potential for researchers to target pathways associated with the gut-microglia connection and overcome CNS diseases.

The Curcumin Analogs 2-Pyridyl Cyclohexanone Induce Apoptosis via Inhibition of the JAK2-STAT3 Pathway in Human Esophageal Squamous Cell Carcinoma Cells.

Multiple modifications to the structure of curcumin have been investigated with an aim to improve its potency and biochemical properties. Previously, we have synthesized a series of curcumin analogs. In the present study, the anticancer effect of 2-pyridyl cyclohexanone, one of the curcumin analogs, on esophageal carcinoma Eca109 and EC9706 cell lines and its molecular mechanisms were investigated. 2-Pyridyl cyclohexanone inhibited the proliferation of Eca109 and EC9706 cells by inducing apoptosis as indicated by morphological changes, membrane phospholipid phosphatidylserine ectropion, caspase 3 activation, and cleavage of poly(ADP-ribose) polymerase. Mechanistic studies indicated that 2-pyridyl cyclohexanone disrupted mitochondrial membrane potential, disturbed the balance of the Bcl-2 family proteins, and triggered apoptosis via the mitochondria-mediated intrinsic pathway. In 2-pyridine cyclohexanone-treated cells, the phosphorylation levels of JAK2 and STAT3 were dose-dependently decreased and p38 and p-ERK signals were notably activated in a dose-dependent manner. Moreover, we found that the addition of S3I-201, a STAT3 inhibitor, led to a decreased expression level of Bcl-2 in Eca109 cells. The chromatin immunoprecipitation assay demonstrated that STAT3 bound to the promoter of Bcl-2 in the Eca109 cells. Furthermore, the mutation of four STAT3 binding sites (-1733/-1723, -1627/-1617, -807/-797, and -134/-124) on the promote of Bcl-2 gene alone attenuated the transcriptional activation of STAT3. In addition, down-regulation of STAT3 resulted in less of transcriptional activity of STAT3 on Bcl-2 expression. These data provide a potential molecular mechanism of the apoptotic induction function of 2-pyridyl cyclohexanone, and emphasize its important roles as a therapeutic agent for esophageal squamous carcinoma.

The IGF2/IGF1R/Nanog Signaling Pathway Regulates the Proliferation of Acute Myeloid Leukemia Stem Cells.

Acute myeloid leukemia is an aggressive disease characterized by clonal proliferation and differentiation into immature hematopoietic cells of dysfunctional myeloid precursors. Accumulating evidence shows that CD34+CD38- leukemia stem cells (LSCs) are responsible for drug resistance, metastasis, and relapse of leukemia. In this study, we found that Nanog, a transcription factor in stem cells, is significantly overexpressed in CD34+ populations from patients with acute myeloid leukemia and in LSCs from leukemia cell lines. Our data demonstrate that the knockdown of Nanog inhibited proliferation and induced cell cycle arrest and cell apoptosis. Moreover, Nanog silencing suppressed the leukemogenesis of LSCs in mice. In addition, we found that these functions of Nanog were regulated by the insulin-like growth factor receptor (IGF1R) signaling pathway. Nanog overexpression rescued the colony formation ability of LSCs treated with picropodophyllin (PPP), an IGF1R inhibitor. By contrast, knockdown of Nanog abolished the effects of IGF2 on the colony formation ability of these LSCs. These findings suggest that the IGF2/IGF1R/Nanog signaling pathway plays a critical role in LSC proliferation.