Protein & Cell

, Volume 8, Issue 5, pp 328–348 | Cite as

Reprogramming cell fates by small molecules

Open Access
Review

ABSTRACT

Reprogramming cell fates towards pluripotent stem cells and other cell types has revolutionized our understanding of cellular plasticity. During the last decade, transcription factors and microRNAs have become powerful reprogramming factors for modulating cell fates. Recently, many efforts are focused on reprogramming cell fates by non-viral and non-integrating chemical approaches. Small molecules not only are useful in generating desired cell types in vitro for various applications, such as disease modeling and cell-based transplantation, but also hold great promise to be further developed as drugs to stimulate patients’ endogenous cells to repair and regenerate in vivo. Here we will focus on chemical approaches for generating induced pluripotent stem cells, neurons, cardiomyocytes, hepatocytes and pancreatic β cells. Significantly, the rapid and exciting advances in cellular reprogramming by small molecules will help us to achieve the long-term goal of curing devastating diseases, injuries, cancers and aging.

Keywords

reprogramming small molecules stem cells cell fates 

INTRODUCTION

Studies of developmental biology have demonstrated that the zygote is totipotent and can gradually differentiate into all tissues and organs of the human body, that is composed by more than 200 types of specialized cells. However, the demonstration of interchanging cell fates and converting one cell type into another by different strategies, such as nuclear transfer or cell fusion, has challenged the traditional cognition of development and regeneration (Gurdon, 1962; Tada et al., 1997). In 2006, Yamanaka and Takahashi reported the development of the induced pluripotent stem cell (iPSC) technology, which can convert somatic cells to iPSCs using four transcription factors (TFs) (Takahashi and Yamanaka, 2006). During the last 10 years, iPSC technology has provided many fundamental insights into the molecular mechanisms of cell fate transition, and already demonstrated promising results in various applications, including disease modeling, drug screening, and cell-based therapy (Takahashi and Yamanaka, 2016). Recently, different combinations of lineage-specific TFs have been screened and applied to generate various cell types, including neurons, cardiomyocytes, hepatocytes, and pancreatic β cells (Huang et al., 2011; Ieda et al., 2010; Vierbuchen et al., 2010; Zhou et al., 2008). However, TF-based reprogramming approaches face many challenges in efficiency, safety, and in vivo delivery. As a novel and promising solution, small molecules are easy to apply and remove, more efficient and amenable to scale up. Small molecules not only are useful in generating desired cell types in vitro for various applications, but also can be further developed as drugs to stimulate patients’ endogenous cells to repair and regenerate in vivo.

In this review, we will focus on making iPSCs, neurons, cardiomyocytes, hepatocytes and pancreatic β cells by small molecules (Fig. 1). Many interesting small molecules have been identified that can significantly promote cellular reprogramming (Table 1). The rapid and exciting advances in cellular reprogramming by small molecules will undoubtedly advance biomedical studies and clinical translation.
Figure 1

Reprogramming cell fates by small molecules. Chemical approaches can be widely applied to manipulate cell fates and states, including pluripotent reprogramming, directed differentiation, and lineage reprogramming. Small molecules not only are useful in generating functional cell types, such as cardiomyocytes, hepatocytes, pancreatic β cells, and neurons, but also can provide a better understanding of the detailed mechanisms underlying specific reprogramming processes

Table 1

Representative small molecules for reprogramming cell fates

Name

Chemical formula

Function

Application

5-Azacytidine (5-aza)

Open image in new window

DNA methyltransferase inhibitor

iPSC reprogramming

5-Aza-2’-deoxycitidine (5-aza-dc)

Open image in new window

DNA methyltransferase inhibitor

iPSC reprogramming

RepSox (E616452)

Open image in new window

ALK5 inhibitor

iPSC reprogramming

Kenpaullone

Open image in new window

GSK3β inhibitor, CDK inhibitor

iPSC reprogramming

BIX01294

Open image in new window

G9a inhibitor

iPSC, iCM, pancreatic β-like cell reprogramming

Bayk8644

Open image in new window

L-type Ca2+ channel agonist

iPSC and pancreatic β-like cell reprogramming

RG108

Open image in new window

DNA methyltransferase inhibitor

iPSC, iNSC,

pancreatic β-like cell reprogramming

CHIR99021

Open image in new window

GSK3β inhibitor

iPSC, iN, iNSC, iCM, iCPC, iHep reprogramming

Tranylcypromine (Parnate)

Open image in new window

Lysine-specific demethylase 1 inhibitor

iPSC,iNSC, iCM reprogramming

Valproic acid (VPA)

Open image in new window

Histone deacetylase inhibitor

iPSC, iNSC, iCM reprogramming

AMI-5

Open image in new window

Protein arginine methyltransferase inhibitor

iPSC reprogramming

A83-01

Open image in new window

TGFβ signaling pathway inhibitor

iPSC, iNSC, iCM, iHep reprogramming

Forskolin

Open image in new window

cAMP signaling activator

iPSC, iN, iCM reprogramming

2-Methyl-5-hydroxytryptamine (2-Me-5-HT)

Open image in new window

5-HT receptor agonist

iPSC reprogramming

D4476

Open image in new window

Casein kinase inhibitor

iPSC reprogramming

3-Deazaneplanocin (DZNep)

Open image in new window

DNA methyltransferase inhibitor

iPSC reprogramming

AM580

Open image in new window

RARα agonist

iPSC reprogramming

EPZ004777

Open image in new window

DOT1L inhibitor

iPSC reprogramming

SGC0946

Open image in new window

DOT1L inhibitor

iPSC reprogramming

Sodium butyrate (NaB)

Open image in new window

Histone deacetylase inhibitor

iPSC and iNSC reprogramming

PS48

Open image in new window

PDK1 activator

iPSC reprogramming

PD0325901

Open image in new window

MAPK/ERK signaling pathway inhibitor

iPSC reprogramming

ISX9

Open image in new window

Activates the expression of endogenous neurogenic transcription factors

iN reprogramming

I-BET151

Open image in new window

BET family protein inhibitor

iN reprogramming

SP600125

Open image in new window

JNK inhibitor

iN and iNSC reprogramming

GO6983

Open image in new window

Pan-PKC inhibitor

iN reprogramming

Dorsomorphin

Open image in new window

BMP signaling inhibitor

iN reprogramming

LDN193189

Open image in new window

BMP signaling inhibitor

iN, iNSC reprogramming

SB431542

Open image in new window

TGFβ signaling pathway inhibitor

iN, iCM, pancreatic β-like cell reprogramming

TTNPB

Open image in new window

Synthetic retinoic acid receptor ligand

iN reprogramming

Thiazovivin (Tzv)

Open image in new window

ROCK inhibitor

iPSC, iN reprogramming

DAPT

Open image in new window

γ-secretase inhibitor

iN reprogramming

Smoothened agonist (SAG)

Open image in new window

Hedgehog signaling activator

iN reprogramming

Purmorphamine (Purmo)

Open image in new window

Hedgehog signaling activator

iN reprogramming

Lysophosphatidic acid (LPA)

Open image in new window

GPCR activator

iNSC reprogramming

Rolipram

Open image in new window

Phosphodiesterase-4 inhibitor

iNSC, iCM reprogramming

Retinoic acid (RA)

Open image in new window

RAR receptor

iNSC reprogramming

Hh-Ag1.5

Open image in new window

Hedgehog signaling activator

iNSC reprogramming

SMER28

Open image in new window

Autophagy enhancer

iNSC reprogramming

Y-27632

Open image in new window

ROCK1 inhibitor

iN, iCM reprogramming

JAK inhibitor I

Open image in new window

JAK inhibitor

iCM reprogramming

LY-364947

Open image in new window

TGFβR-I inhibitor

iCM reprogramming

SD-208

Open image in new window

TGFβR-I inhibitor

iCM reprogramming

GW788388

Open image in new window

ALK5 inhibitor

iCM reprogramming

ICARIIN

Open image in new window

Exhibits a variety of pharmacological actions

iCM reprogramming

PD169316

Open image in new window

p38 inhibitor

iCM reprogramming

AS8351

Open image in new window

KDM5B inhibitor

iCM reprogramming

SC1

Open image in new window

ERK1, RasGAP inhibitor

iCM reprogramming

OAC2

Open image in new window

Oct4 activator

iCM reprogramming

SU16F

Open image in new window

PDGFR inhibitor

iCM reprogramming

JNJ10198409

Open image in new window

PDGFR inhibitor

iCM reprogramming

SU5402

Open image in new window

FGFR, VEGFR, and PDGFR inhibitor

iCPC reprogramming

Compound E

Open image in new window

Notch signaling inhibitor

iHep, pancreatic β-like cell reprogramming

Nicotinamide

Open image in new window

Coenzyme, cofactor

Pancreatic β-cell reprogramming

5’-N-ethylcarboxamidoadeno-sine (NECA)

Open image in new window

Adenosine receptor agonist

Pancreatic β-like cell reprogramming

BRD7552

Open image in new window

PDX1 expression inducer

Increases insulin expression

Vitamin C

Open image in new window

Cofactor of epigenetic modulators

Increases reprogramming efficiency

PLURIPOTENT REPROGRAMMING

In 2006, the first generation of iPSCs were derived by using four transcription factors (Oct4, Sox2, Klf4, and c-Myc, termed OSKM) (Takahashi and Yamanaka, 2006). During the last 10 years, substantial efforts have been made to improve the efficiency and safety of iPSC reprogramming by non-integrating viral vectors, synthetic RNAs, recombinant proteins and small molecules (Gonzalez et al., 2011). Particularly, screenings have been applied to identify chemical compounds that can replace individual or multiple Yamanaka factors. Ichida et al. carried out a high-content chemical screening and identified RepSox (also named as E616452, an ALK5 inhibitor) as a chemical replacer of Sox2 (Ichida et al., 2009). The authors suggested that RepSox promoted the completion of reprogramming through induction of Nanog. In addition, RepSox possibly promoted mesenchymal-to-epithelial transition (MET) during the early phase of reprogramming (Li et al., 2010). Using a Nanog-Luciferase reporter as readout, Kenpaullone was found to functionally replace Klf4 in the presence of OSM (Lyssiotis et al., 2009). Shi et al. found that the combination of BIX01294 and Bayk8644 or BIX01294 and RG108 enabled OK-induced reprogramming of mouse fibroblasts, and Li et al. further reported successful OK-reprogramming of human somatic cells by using CHIR99021 and Tranylcypromine (also named Parnate) (Li et al., 2009; Shi et al., 2008). Huangfu et al. reported that Valproic acid (VPA) could promote OS-induced reprogramming of human fibroblasts (Huangfu et al., 2008). Taken together, these studies demonstrated that epigenetic regulators play important roles in iPSC reprogramming. Subsequently, several groups have focused on Oct4-only reprogramming by employing different chemical cocktails. Chen et al. found that bone morphogenetic proteins (BMPs) could support efficient mouse Oct4-alone reprogramming partly through the promotion of MET (Chen et al., 2011). Interestingly, Yuan et al. reported that AMI-5 and A83-01 enabled Oct4-induced reprogramming of mouse fibroblasts. Remarkably, these Oct4-only iPSCs were truly pluripotent and could give rise to live-born pups by tetraploid complementation assay (Yuan et al., 2011). Furthermore, Li et al. identified a specific chemical combination, including VPA, CHIR99021, E616452 and Tranylcypromine, which was sufficient to reprogramming mouse fibroblasts to iPSCs with Oct4 alone (Li et al., 2011). Subsequently, some interesting chemical substitutes of Oct4 in mouse iPS-reprogramming were identified, including Forskolin, D4476, and 2-Methyl-5-hydroxytryptamine (2-Me-5-HT) (Hou et al., 2013).

All chemical induced pluripotent reprogramming can provide a promising paradigm for cell fate transitions. In 2013, Hou et al. demonstrated successful reprogramming of mouse cells into iPS cells for the first time by using a novel cocktail of seven small molecules, including VPA, CHIR99021, E616452, Tranylcypromine, Forskolin, 3-deazaneplanocin A (DZNep), and TTNPB (Hou et al., 2013). Further refinement of this all-chemical reprogramming led a 1000-fold greater efficiency with additional small molecules, including AM580, EPZ004777, SGC0946, and 5-aza-2-deoxycitidine (5-aza-dC) (Zhao et al., 2015b). Interestingly, chemical induced pluripotent reprogramming process required the early formation of extra-embryonic endoderm (XEN)-like cells and a late transition from XEN-like cells to iPSCs, which fundamentally distinct from the pathway of TF-induced pluripotent reprogramming. However, it is still largely unknown how small molecules activate the endogenous gene regulation network, and gradually enable cell fate transitions. The remaining question is obvious: how to achieve chemical induced pluripotent reprogramming of human cells? We previously made some progress and reported that a chemical cocktail of Sodium butyrate (NaB), Parnate, PS48, CHIR99021, A83-01 and PD0325901 enabled OCT4-induced reprogramming of human primary somatic cells (Zhu et al., 2010). These human OCT4-only iPSCs were pluripotent and could give rise to cells of all three germ layers both in vitro and in vivo. Further mechanistic studies demonstrated that a metabolic switch from oxidative phosphorylation to glycolysis is an important step in iPSC reprogramming. More efforts are required to identify chemical substitutes for OCT4 in human cell reprogramming. The demonstration of human pluripotent reprogramming by an all-chemical approach will greatly benefit the study of stem cell biology and regenerative medicine.

NEURAL REPROGRAMMING

An increasing number of people suffer from neurodegenerative disorders, such as Alzheimer’s disease and Parkinson’s disease, as life expectancy further extends. It is not feasible to obtain sufficient amounts of patient specific neural cells for disease modeling and drug development. In the last few years, great progress has been made using TF-based direct reprogramming towards induced neurons (iNs) or induced neural stem cells (iNSCs).

In 2010, Vierbuchen et al. firstly reported that mesodermal mouse fibroblasts could be rapidly and efficiently reprogrammed into ectodermal iNs by using three TFs: Brn2, Ascl1, and Myt1 l (BAM) (Vierbuchen et al., 2010). Subsequently, BAM plus NEUROD1 was applied for neuronal reprogramming of human cells (Pang et al., 2011). So far, a number of groups have generated different subtypes of neurons by combining BAM with specific TFs that play an important role in the development of specific neuron subtypes (Caiazzo et al., 2011; Kim, 2011; Pang et al., 2011; Son et al., 2011). Besides TFs, microRNAs, such as miR-9 and miR-124, can also facilitate neuronal reprogramming (Ambasudhan et al., 2011; Yang et al., 2016; Yoo et al., 2011). Obviously, it is desirable to develop non-viral and non-integrative iN reprogramming approaches by small molecules. In 2015, two simultaneous articles reported the successful generation of iN from mouse and human fibroblasts via chemical-only approaches (Hu et al., 2015; Li et al., 2015). Li et al. showed that mouse fibroblasts could be converted into neurons by using an optimal cocktail of four small molecules (Forskolin, ISX9, CHIR99021, and I-BET151, termed FICB) (Li et al., 2015). The authors suggested that I-BET151, a BET family protein inhibitor, suppressed the fibroblast-specific program, and ISX9 activated the expression of endogenous neurogenic transcription factors, which synergistically promoted neuronal conversion (Li et al., 2015). Hu et al. demonstrated the generation of neurons from human fibroblasts using a small molecule cocktail (VPA, CHIR99021, RepSox, Forskolin, SP600125, GO6983, Y-27631, and Dorsomorphin) (Hu et al., 2015). Furthermore, via this chemical cocktail, fibroblasts from familial Alzheimer’s disease patients could also be reprogrammed to iNs. Such cells can be used for in vitro disease modeling and drug screenings (Hu et al., 2015). Subsequently, Zhang et al. reported that sequential addition of a cocktail of small molecules (LDN193189, SB431542, TTNPB, Thiazovivin (Tzv), CHIR99021, VPA, DAPT, Smoothened agonist (SAG), and Purmorphamine) can reprogram human astrocytes into functional neurons (Zhang et al., 2015). Mechanistically, these small molecules inhibited glial but activated neuronal signaling pathways through epigenetic and transcriptional modulation. Remarkably, these human iNs were functional and could survive more than 5 months under cell culture conditions.

Compared with neurons, expandable and multipotent iNSCs are desirable for downstream applications, like disease modeling and drug screening. In the past few years, many groups reported the generation of iNSCs using neural lineage-specific TFs. These iNSCs are multipotent and can differentiate into functional neurons, astrocytes, and oligodendrocytes both in vitro and in vivo (Ring et al., 2012; Zhou and Tripathi, 2012). Recently, we achieved both mouse and human iNSC reprogramming by a cell-activation signaling-directed (CASD) strategy (Kim et al., 2011; Zhu et al., 2015). The CASD strategy uses transient exposure of somatic cells to reprogramming factors (Oct4, Sox2, Klf4, and c-Myc) in conjunction with soluble lineage-specific signals to reprogram cells into other cell types, such as iNSCs. Several interesting small molecules could promote OCT4-mediated iNSC reprogramming process, including A83-01, CHIR99021, NaB, Lysophosphatidic acid (LPA), Rolipram and SP600125 (Zhu et al., 2014a). Furthermore, similar to iN reprogramming, there are also great advances in iNSC reprogramming by using small molecules alone. In 2014, Cheng and colleagues used three small molecules VPA, CHIR99021, and RepSox to derive iNPCs from somatic cells (Cheng et al., 2014). More recently, Zhang et al. achieved more efficient mouse iNSC reprogramming by using a cocktail of nine components (CHIR99021, LDN193189, A83-01, Retinoic acid (RA), Hh-Ag1.5, RG108, Parnate, SMER28, and bFGF) (Zhang et al., 2016a). They provided definitive evidence that these iNSCs could be reprogrammed from fibroblasts using a genetic lineage-tracing system. Interestingly, further mechanistic studies uncovered that these small molecules could gradually and specifically activate key neurogenic regulators, such as Sox2, and then facilitated the neural cell fate transition.

Direct in vivo reprogramming will provide a perspective for cell-based clinical regenerative therapy (Chen et al., 2015; Li and Chen, 2016). Glial cells are the most abundant cells in adult brains and several groups have reported the successful TF-based reprogramming of glial cells to neurons or iNPCs. Niu et al. found that delivery of Sox2 could reprogram endogenous astrocytes to proliferating neuroblasts and these neuroblasts further differentiated to functional neurons that integrated into neural networks in the brain (Niu et al., 2013). Guo et al. demonstrated that cortical glial cells activated by injury or disease could be reprogrammed by NeuroD1 in vivo (Guo et al., 2014). The further application of knowledge learned from in vitro chemical screening and ambitious in vivo chemical screening will undoubtedly advance this field.

CARDIAC REPROGRAMMING

The adult mammalian heart possesses little regenerative capacity following injury. Cardiac fibroblasts account for a majority of cells in the heart, and cardiac reprogramming holds great potentials. In 2010, Ieda et al. reported that postnatal cardiac fibroblasts could be directly reprogrammed into induced cardiomyocyte-like cells (iCMs) by transfection with a combination of three TFs (Gata4, Mef2c, Tbx5, termed GMT) (Ieda et al., 2010). Lineage-tracing experiments showed that the cardiac reprogramming with GMT was a direct conversion process. Subsequently, other groups showed that addition of TFs such as Hand2 and Nkx2.5 to GMT promoted the reprogramming efficiency or maturation of iCMs (Addis and Epstein, 2013; Ifkovits et al., 2014; Song et al., 2012). Additionally,miRNAs, such as miR-1 and miR-133, also play important roles in cardiac reprogramming (Ieda, 2016; Jayawardena et al., 2012; Muraoka et al., 2014; Nam et al., 2013; Zhao et al., 2015a). Although the efficiency of cardiac reprogramming has been improved in recent years, the molecular mechanisms of this process are largely unknown. More recently, Zhou et al. carried out a small-scale functional screening and identified that loss of Bmi1 significantly promoted mouse cardiac reprogramming. Mechanistically, Bmi1 blocked cardiac reprogramming through direct interactions with the regulatory regions of many cardiogenic genes (Zhou et al., 2016). Compared with mouse cardiac reprogramming, human cardiac reprogramming is much more challenging to be achieved. Fu et al. showed that GMT plus ESRRG, MESP1, Myocardin, and ZFPM2 could reprogram human fibroblasts to iCMs (Fu et al., 2013). Nam et al. demonstrated that human fibroblasts could be converted to iCMs by introduction of GATA4, HAND2, TBX5, Myocardin, miR-1, and miR-133 under long-term culture (Nam et al., 2013). Wada et al. found that GMT plus MESP1 and Myocardin could reprogram human fibroblasts to iCMs (Rie et al., 2013). Such TF-reprogrammed iCMs exhibited sarcomere formation, calcium transients, and action potentials, but could not contract spontaneously and robustly. These results indicated that further improvements are still required for complete human cardiac reprogramming.

Small molecules can potentially replace TFs and provide a novel approach for cardiac reprogramming. Interestingly, the addition of TGF-β inhibitor SB431542 or A83-01 improved the efficiency of iCM generation (Fu et al., 2013; Zhao et al., 2015a). Other small molecules, including Y-27632, JAK inhibitor I, LY-364947, SD-208 and GW788388, did also enhance cardiac reprogramming (Lalit et al., 2016; Zhao et al., 2015a, b). Tamakawa et al. showed that adding fibroblast growth factor (FGF) 2, FGF10, and vascular endothelial growth factor (VEGF) to GMT enhanced cardiac reprogramming about 100-fold compared with GMT only (Yamakawa et al., 2015). Wang et al. identified a small molecule cocktail, including CHIR99021, SB431542, Parnate, and Forskolin, that enabled efficient conversion of mouse fibroblasts into iCMs with Oct4 alone employing the CASD paradigm. These iCMs spontaneously contracted and exhibited a ventricular phenotype (Wang et al., 2014). Recently, there have been significant achievements in cardiac reprogramming by using small molecules alone. Fu et al. showed that a chemical cocktail (CHIR99021, RepSox, Forskolin, and VPA) could induce beating clusters of cardiac cells from mouse fibroblasts. Other small molecules, including ICARIIN, PD169316 and Rolipram also increased the efficiency of cardiac reprogramming (Fu et al., 2015). Nan et al. showed that human iCMs could be generated from fibroblasts by a specific set of small molecules (Cao et al., 2016). This combination of nine compounds was termed 9C, including CHIR99021, A83-01, BIX01294, AS8351, SC1, Y-27632, OAC2, SU16F, and JNJ10198409. Remarkably, at both transcriptome and epigenetic levels, these human iCMs resembled human cardiomyocytes. The authors observed enrichment of H3K4me3 as well as H3K27ac, and decrease of H3K27me3 on a cohort of heart developmental genes during 9C-induced cardiac reprogramming. Mechanistically, 9C treatment epigenetically and transcriptionally activated key cardiac developmental genes and facilitated the cardiac fate transition. Interestingly, unlike human iCMs from TF-based reprogramming, these human 9C-induced iCMs uniformly contracted. More efforts are required to apply this chemical cardiac reprogramming approach to patient cells.

Expandable and multipotent induced cardiac progenitor cells (iCPCs) are more suitable for basic studies and translational applications. Islas et al. found that human dermal fibroblasts could be converted into cardiac progenitors by ETS2 and MESP1 overexpression (Islas et al., 2012). After a screening of multiple cardiac TFs, Lalit et al. reported that a combination of Mesp1, Tbx5, Gata4, Nkx2.5, and Baf60c could reprogram mouse fibroblasts into iCPCs (Lalit et al., 2016). Meanwhile, Zhang et al. demonstrated that fibroblasts could be induced into expandable iCPCs by the CASD lineage conversion strategy and these iCPCs could be further long-term expanded using a combination of growth factors and small molecules (BMP4, Activin A, CHIR99021 and SU5402) (Zhang et al., 2016b). The activation of Wnt signaling by CHIR99021 not only promoted iCPC proliferation, but also inhibited further differentiation of iCPCs into cardiomyocytes. In addition, treating iCPC with Wnt inhibitor IWP2 facilitated even more efficient cardiac differentiation. Reprogramming mouse and human fibroblasts into expandable cardiac progenitors by small molecules alone could likely be achieved with further screening and modifications in the future.

Cardiac fibroblasts account for a majority of cells in the heart and represent a potential cell source for cardiac reprogramming. Direct in vivo cardiac reprogramming has already demonstrated some promising results. Qian et al. converted mouse resident non-myocytes into iCMs in vivo by local delivery of GMT after coronary ligation (Qian et al., 2012). Song et al. showed that GMT plus Hand2 could cooperatively reprogram dividing non-cardiomyocytes into functional iCMs in situ (Song et al., 2012). These cardiac reprogramming TF-delivered hearts had decreased infarct size and better cardiac function. Therefore, reprogramming cardiac fibroblasts to iCMs in their native environment by chemical compounds will hold great potentials in cardiac regenerative therapies.

HEPATIC REPROGRAMMING

The liver is a fascinating organ that can regenerate itself; but unfortunately, this remarkable capacity is lost in chronic liver diseases. Currently, there are over 600 million people with liver diseases and 1 million deaths per year worldwide. Because the limitation of organ donors, cell-based therapy may represent an attractive approach for treating life-threatening liver failure, if a sufficient amount of functional hepatocytes can be generated from easily accessible cell sources. Many research groups focus their efforts on the generation of induced hepatocyte-like cells (iHeps) via reprogramming approaches. In 2011, Huang et al. showed that functional iHep cells could be converted from p19 null mouse fibroblasts via transduction of Gata4, Hnf1α, and Foxa3 (Huang et al., 2011). Moreover, these iHep cells restored liver functions in vivo and rescued the recipients from death, which provided a promising proof of principle example for the use in regenerative medicine. Meanwhile, Sekiya et al. converted mouse embryonic and adult fibroblasts into iHep cells using a combination of Hnf4α plus Foxa1, Foxa2 or Foxa3 (Sekiya and Suzuki, 2011). Remarkably, after overexpressing Hnf1β and Foxa3, mouse embryonic fibroblasts were converted to induced hepatic stem cells, which had the capability of long-term expansion and bipotential differentiation to hepatocytes and cholangiocytes (Yu et al., 2013). Subsequently, human fibroblasts were successfully reprogrammed into iHep cells by different combinations of TFs (Du et al., 2014; Huang et al., 2014). Huang et al. generated human iHeps from fibroblasts by introduction of FOXA3, HNF1α, and HNF4α (Huang et al., 2014). Meanwhile, Du et al. showed that human fibroblasts could be reprogrammed to functional iHeps by overexpressing HNF1α, HNF4α, HNF6, ATF5, PROX1, and CEBPα (Du et al., 2014). These human iHeps expressed liver-specific markers, and could integrate into mouse liver and demonstrated many metabolic functions of the liver.

Recently, the non-viral and non-integrating transduction systems, including synthetic RNAs, recombinant proteins, and episomal plasmids, become more attractive (Fusaki et al., 2009; Kim et al., 2009, 2015; Warren et al., 2010; Zhou and Freed, 2009). For example, Simeonov et al. used a method of repeating transfection with synthetic modified mRNAs encoding hepatic reprogramming TFs and successfully induced iHep cells from human fibroblasts (Simeonov and Uppal, 2014). Kim et al. applied an oriP/EBNA1-based episomal system to deliver a set of transcription factors, including Gata4, Hnf1α, and Foxa3, to convert fibroblasts into iHep cells (Kim et al., 2015). Small molecules also played a critical role in iHep reprogramming process (Lim et al., 2016; Pournasr et al., 2015). We previously generated induced endodermal progenitor cells by the CASD lineage conversion strategy, and greatly expanded these endodermal progenitor cells by a chemically defined condition, consisting of EGF, bFGF, A83-01, and CHIR99021. Further treatment with soluble growth factors and small molecules, such as A83-01 and Compound E, these induced endodermal progenitor cells efficiently differentiated into functional hepatocytes. After transplantation, human iHeps integrated into immunodeficient mouse livers, expanded extensively, and acquired mature hepatocyte functions (Zhu et al., 2014b). Moreover, Lim et al. reported that direct conversion into iHeps is a stepwise transition involving the sequential erasure of somatic memory, MET transition, and induction of hepatic cell fate (Lim et al., 2016). Through chemical screening, they found that CHIR99021 and A83-01 facilitated one factor Hnf1α-induced hepatic reprogramming. Mechanistically, these small molecules facilitated the robust iHep generation through promoting MET process. In the future, utilizing all-chemical method to induce iHep reprogramming will be a promising way to promote the development of this field.

More recently, in vivo hepatic reprogramming was fulfilled by injection of hepatic reprogramming factors in situ (Rezvani et al., 2016; Song et al., 2016). Rezvani et al. developed in vivo reprogramming of myofibroblasts into hepatocytes using adeno-associated virus (AAV) vectors expressing hepatic transcription factors: Foxa1, Foxa2, Foxa3, Gata4, Hnf1a, and Hnf4a. These iHeps converted from myofibroblsts were functional and reduced liver fibrosis (Rezvani et al., 2016). Song et al. showed that overexpression of FOXA3, GATA4, HNF1A, and HNF4A could convert mouse myofibroblasts into iHeps cells in fibrotic mouse livers and also reduced liver fibrosis (Song et al., 2016). More efforts are required to develop better delivery methods and improve the conversion efficiency and cell maturity. Obviously, in vivo hepatic reprogramming by small molecules will facilitate the translation of the iHep technology into clinical treatment of chronic liver diseases.

PANCREATIC REPROGRAMMING

Diabetes mellitus represents a global health epidemic and affects more than 300 million people worldwide according to the International Diabetes Federation. Diabetes is caused and developed as a deficiency and/or dysfunction of pancreatic β cells. A key method to study diabetes and treat patients is to obtain unlimited numbers of functional pancreatic β cells. Reprogramming other types of somatic cells to pancreatic β cells is under active investigation. In 2005, Minammi et al. reported that adult mouse pancreatic exocrine cells could be converted into insulin-producing cells in vitro by suspension culture with EGF and nicotinamide (Minami et al., 2005). In 2008, Zhou et al. reprogrammed adult pancreatic exocrine cells to β-like cells in vivo by introducing three TFs: Pdx1, Ngn3, and Mafa (Zhou et al., 2008). Then, Li et al. developed an improved method for in vivo pancreatic conversion (Li et al., 2014b). These induced pancreatic β-like cells formed the islet-like structures and could persist in vivo for more than one year (Li et al., 2014a). Additionally, Chen et al. reported that transient intestinal expression of Pdx1, Ngn3, and Mafa promoted rapid conversion of intestinal crypt cells into endocrine cells (Chen et al., 2014). Besides mouse cells, human pancreatic ductal cells could also be converted into pancreatic β-like cells by overexpression of PDX1, NGN3, MAFA, and PAX6 (Lee et al., 2013). These converted pancreatic β-like cells could secrete insulin in response to high concentration of glucose. More interestingly, Ariyachet et al. reported that cells of the antral stomach were ideal cell source for pancreatic reprogramming (Ariyachet et al., 2016). These reprogrammed insulin+ β-like cells performed molecularly and functionally similar to bona fide pancreatic β cells. These results suggested that the initial cell type can affect the reprogramming efficiency and induced cell phenotype and its functional properties.

Introducing exogenous genetic material and altering the genome raise safety concerns (Barrilleaux and Knoepfler, 2011). Therefore, using small molecules to active endogenous TFs is an attractive method for cellular reprogramming. By high-throughput screening, BRD7552 was shown to be an inducer of PDX1 expression (Yuan et al., 2013). In another report, DNA methyltransferase inhibitor 5-aza-dC was capable of increasing the expression of Ngn3 in PANC-1 cells (Lefebvre et al., 2010). In our previous study, we found that several specific small molecules could enhance the endodermal conversion process, including CHIR99021, NaB, Parnate, RG108, and 5’-N-ethylcarboxamidoadenosine, and promote pancreatic β-like cell differentiation and maturation, including Compound E, Vitamin C, and Bayk8644 (Zhu et al., 2016). More recently, Wang et al. described the derivation of human induced endodermal progenitor cells from gastrointestinal epithelial cells using a cocktail of defined small molecules, including Bayk8644, BIX01294, RG108, and SB431542, along with support from tissue-specific mesenchymal feeders (Wang et al., 2016). Induced endodermal progenitor cells could subsequently differentiate into more specified cell types, including pancreatic β-like cells. However, the therapeutic potentials of these pancreatic β-like cells have not been investigated so far. Obviously, using small molecules to replace master TFs as well as promote reprogramming to mature pancreatic β cells is a promising therapy for treating diabetes. However, transferring these findings for efficient in vivo pancreatic reprogramming, preferably be an all-chemical approach is waiting to be achieved in the future.

PERSPECTIVE

Cell fate transition and regeneration have fascinated biologists for centuries. After the discovery of iPSC technology, a lot of progress has been made in TF-mediated cellular reprogramming into many different defined cell types. Compared with the introduction of TFs by viral methods, small molecules have several advantages, can enhance reprogramming efficiency and improve the quality of the reprogrammed cells. Currently, identification of novel and effective small molecules for cellular reprogramming remains very labor intense. New technologies, such as high-throughput screening (HTS2) platform (Li et al., 2012) and reporter systems generated by CRISPR tools (Hockemeyer and Jaenisch, 2016), can be applied to this field and will accelerate the discovery process. In addition, the molecular mechanisms of most reprogramming processes are largely unknown, and how TFs or small molecules mediate cell fate transitions is still unclear. New technologies, such as single-cell analysis (Wen and Tang, 2016) and CRISPR-based genome-wide screening (Shalem et al., 2014), can provide new insights and help us to better understand the underlying molecular mechanisms. A better understanding of the detailed mechanisms during reprogramming processes can also help to improve the reprogramming efficiency and acquire mature cells with complete functionality for disease modeling, drug development, and cell-based transplantation (Fig. 2). Optimistically, future studies in cellular reprogramming by small molecules will overcome current hurdles, generate new discoveries, and benefit human health.
Figure 2

Potential applications of cellular reprogramming. Cellular reprogramming can provide a large number of functional cells, which can be used for cell-based transplantation and high-throughput chemical screenings. This technology will help develop drugs to stimulate patients’ endogenous cells to repair and regenerate in vivo in the near future. Cellular reprogramming by only small molecules will significantly advance biomedical studies and clinical applications, and realize the long-term goal of curing degenerative diseases, injuries, and aging

Notes

ACKNOWLEDGEMENTS

We would like to thank Dr. Wanguo Wei at Shanghai Advanced Research Institute, Chinese Academy of Sciences and Dr. Holger A. Russ at the Barbara Davis Center for Childhood Diabetes at the University of Colorado for their generous help. This work was funded by the national key research and development programme of China (2016YFC1305300) and the outstanding youth fund of Zhejiang Province (R17C120002). S.Z. is a scholar of the National 100 Young Talents Program. We apologize to those whose findings are relevant but not cited in this review due to space limitation.

ABBREVIATIONS

2-Me-5-HT, 2-Methyl-5-hydroxytryptamine; AAV, adeno-associated virus; BAM, Brn2, Ascl1, and Myt1 l; BMP, bone morphogenetic protein; CASD, cell-activation signaling-directed; FGF, fibroblast growth factor; iCM, induced cardiomyocyte-like cell; iCPC, induced cardiac progenitor cell; iN, induced neuron; iNSC, induced neural stem cell; iPSC, induced pluripotent stem cell; LPA, Lysophosphatidic acid; NaB, Sodium butyrate; RA, Retinoic acid; SAG, Smoothened agonist; TF, transcription factor; Tzv, Thiazovivin; VEGF, vascular endothelial growth factor; XEN, extra-embryonic endoderm.

COMPLIANCE WITH ETHICS GUIDELINES

Xiaojie Ma, Linghao Kong, and Saiyong Zhu declare that they have no conflict of interest.

REFERENCES

  1. Addis RC, Epstein JA (2013) Induced regeneration–the progress and promise of direct reprogramming for heart repair. Nat Med 19:829–836CrossRefPubMedGoogle Scholar
  2. Ambasudhan R, Talantova M, Coleman R, Yuan X, Zhu S, Lipton SA, Ding S (2011) Direct reprogramming of adult human fibroblasts to functional neurons under defined conditions. Cell Stem Cell 9:113–118CrossRefPubMedPubMedCentralGoogle Scholar
  3. Ariyachet C, Tovaglieri A, Xiang G, Lu J, Shah MS, Richmond CA, Zhou Q (2016) Reprogrammed stomach tissue as a renewable source of functional beta cells for blood glucose regulation. Cell Stem Cell 18:410–421CrossRefPubMedPubMedCentralGoogle Scholar
  4. Barrilleaux B, Knoepfler P (2011) Transduction of human cells with polymer-complexed ecotropic lentivirus for enhanced biosafety. J Vis Exp 53:1–7Google Scholar
  5. Caiazzo M, Dell’Anno MT, Dvoretskova E, Lazarevic D, Taverna S, Leo D, Broccoli V (2011) Direct generation of functional dopaminergic neurons from mouse and human fibroblasts. Nature 476:224–227CrossRefPubMedGoogle Scholar
  6. Cao N, Huang Y, Zheng J, Spencer CI, Zhang Y, Fu J, Nie B, Xie M, Zhang M, Wang H, Ma T, Xu T, Shi G, Srivastava D, Ding S (2016) Conversion of human fibroblasts into functional cardiomyocytes by small molecules. Science 352:1216–1220CrossRefPubMedGoogle Scholar
  7. Chen J, Liu J, Yang J, Chen Y, Chen J, Ni S, Song H, Zeng L, Ding K, Pei D (2011) BMPs functionally replace Klf4 and support efficient reprogramming of mouse fibroblasts by Oct4 alone. Cell Res 21:205–212CrossRefPubMedGoogle Scholar
  8. Chen Y, Finkbeiner SR, Weinblatt D, Emmett MJ, Tameire F, Yousefi M, Yang C, Maehr R, Zhou Q, Shemer R, Dor Y, Li C, Spence JR, Stanger BZ (2014) De novo formation of insulin-producing “neo-beta cell islets” from intestinal crypts. Cell Rep 6:1046–1058CrossRefPubMedPubMedCentralGoogle Scholar
  9. Chen G, Wernig M, Berninger B, Nakafuku M, Parmar M, Zhang CL (2015) In vivo reprogramming for brain and spinal cord repair. eNeuro 2:e0106-15CrossRefGoogle Scholar
  10. Cheng L, Hu W, Qiu B, Zhao J, Yu Y, Guan W, Wang M, Yang W, Pei G (2014) Generation of neural progenitor cells by chemical cocktails and hypoxia. Cell Res 24:665–679CrossRefPubMedPubMedCentralGoogle Scholar
  11. Du Y, Wang J, Jia J, Song N, Xiang C, Xu J, Hou Z, Su X, Liu B, Jiang T, Zhao D, Sun Y, Shu J, Guo Q, Yin M, Sun D, Lu S, Shi Y, Deng H (2014) Human hepatocytes with drug metabolic function induced from fibroblasts by lineage reprogramming. Cell Stem Cell 14:394–403CrossRefPubMedGoogle Scholar
  12. Fu J, Stone NR, Liu L, Spencer CI, Qian L, Hayashi Y, Delgado-Olguin P, Ding S, Bruneau BG, Srivastava D (2013) Direct reprogramming of human fibroblasts toward a cardiomyocyte-like state. Stem Cell Reports 1:235–247CrossRefPubMedPubMedCentralGoogle Scholar
  13. Fu Y, Huang C, Xu X, Gu H, Ye Y, Jiang C, Qiu Z, Xie X (2015) Direct reprogramming of mouse fibroblasts into cardiomyocytes with chemical cocktails. Cell Res 25:1013–1024CrossRefPubMedPubMedCentralGoogle Scholar
  14. Fusaki N, Ban H, Nishiyama A, Saeki K, Hasegawa M (2009) Efficient induction of transgene-free human pluripotent stem cells using a vector based on Sendai virus, an RNA virus that does not integrate into the host genome. Proce Jpn Acad Ser B 85:348–362CrossRefGoogle Scholar
  15. Gonzalez F, Boue S, Izpisua Belmonte JC (2011) Methods for making induced pluripotent stem cells: reprogramming a la carte. Nat Rev Genet 12:231–242CrossRefPubMedGoogle Scholar
  16. Guo Z, Zhang L, Wu Z, Chen Y, Wang F, Chen G (2014) In vivo direct reprogramming of reactive glial cells into functional neurons after brain injury and in an Alzheimer’s disease model. Cell Stem Cell 14:188–202CrossRefPubMedGoogle Scholar
  17. Gurdon JB (1962) The developmental capacity of nuclei taken from intestinal epithelium cells of feeding tadpoles. J Embryol Exp Morphol 10:622–640PubMedGoogle Scholar
  18. Hockemeyer D, Jaenisch R (2016) Induced pluripotent stem cells meet genome editing. Cell Stem Cell 18:573–586CrossRefPubMedPubMedCentralGoogle Scholar
  19. Hou P, Li Y, Zhang X, Liu C, Guan J, Li H, Zhao T, Ye J, Yang W, Liu K, Ge J, Xu J, Zhang Q, Zhao Y, Deng H (2013) Pluripotent stem cells induced from mouse somatic cells by small-molecule compounds. Science 341:651–654CrossRefPubMedGoogle Scholar
  20. Hu W, Qiu B, Guan W, Wang Q, Wang M, Li W, Gao L, Shen L, Huang Y, Xie G, Zhao H, Jin Y, Tang B, Yu Y, Zhao J, Pei G (2015) Direct conversion of normal and Alzheimer’s disease human fibroblasts into neuronal cells by small molecules. Cell Stem Cell 17:204–212CrossRefPubMedGoogle Scholar
  21. Huang P, He Z, Ji S, Sun H, Xiang D, Liu C, Hu Y, Wang X, Hui L (2011) Induction of functional hepatocyte-like cells from mouse fibroblasts by defined factors. Nature 475:386–389CrossRefPubMedGoogle Scholar
  22. Huang P, Zhang L, Gao Y, He Z, Yao D, Wu Z, Cen J, Chen X, Liu C, Hu Y, Lai D, Hu Z, Chen L, Zhang Y, Cheng X, Ma X, Pan G, Wang X, Hui L (2014) Direct reprogramming of human fibroblasts to functional and expandable hepatocytes. Cell Stem Cell 14:370–384CrossRefPubMedGoogle Scholar
  23. Huangfu D, Maehr R, Guo W, Eijkelenboom A, Snitow M, Chen AE, Melton DA (2008) Induction of pluripotent stem cells by defined factors is greatly improved by small-molecule compounds. Nat Biotechnol 26:795–797CrossRefPubMedGoogle Scholar
  24. Ichida JK, Blanchard J, Lam K, Son EY, Chung JE, Egli D, Loh KM, Carter AC, Di Giorgio FP, Koszka K, Huangfu D, Akutsu H, Liu DR, Rubin LL, Eggan K (2009) A small-molecule inhibitor of tgf-Beta signaling replaces sox2 in reprogramming by inducing nanog. Cell Stem Cell 5:491–503CrossRefPubMedPubMedCentralGoogle Scholar
  25. Ieda M (2016) Heart development, diseases, and regeneration- new approaches from innervation, fibroblasts, and reprogramming. Circ J 80:2081–2088CrossRefPubMedGoogle Scholar
  26. Ieda M, Fu J, Delgado-Olguin P, Vedantham V, Hayashi Y, Bruneau BG, Srivastava D (2010) Direct reprogramming of fibroblasts into functional cardiomyocytes by defined factors. Cell 142:375–386CrossRefPubMedPubMedCentralGoogle Scholar
  27. Ifkovits JL, Addis RC, Epstein JA, Gearhart JD (2014) Inhibition of TGFbeta signaling increases direct conversion of fibroblasts to induced cardiomyocytes. PLoS One 9:e89678CrossRefPubMedPubMedCentralGoogle Scholar
  28. Islas JF, Liu Y, Weng K-C, Robertson MJ, Zhang S, Prejusa A, Harger J, Tikhomirova D, Chopra M, Iyer D, Mercola M, Oshima RG, Willerson JT, Potaman VN, Schwartz RJ (2012) Transcription factors ETS2 and MESP1 transdifferentiate human dermal fibroblasts into cardiac progenitors. Proc Natl Acad Sci USA 109:13016–13021CrossRefPubMedPubMedCentralGoogle Scholar
  29. Jayawardena TM, Egemnazarov B, Finch EA, Zhang L, Payne JA, Pandya K, Zhang Z, Rosenberg P, Mirotsou M, Dzau VJ (2012) MicroRNA-mediated in vitro and in vivo direct reprogramming of cardiac fibroblasts to cardiomyocytes. Circ Res 110:1465–1473CrossRefPubMedPubMedCentralGoogle Scholar
  30. Kim KS (2011) Converting human skin cells to neurons: a new tool to study and treat brain disorders? Cell Stem Cell 9:179–181CrossRefPubMedGoogle Scholar
  31. Kim D, Kim CH, Moon JI, Chung YG, Chang MY, Han BS, Ko S, Yang E, Cha KY, Lanza R, Kim KS (2009) Generation of human induced pluripotent stem cells by direct delivery of reprogramming proteins. Cell Stem Cell 4:472–476CrossRefPubMedPubMedCentralGoogle Scholar
  32. Kim J, Efe JA, Zhu S, Talantova M, Yuan X, Wang S, Lipton SA, Zhang K, Ding S (2011) Direct reprogramming of mouse fibroblasts to neural progenitors. Proc Natl Acad Sci USA 108:7838–7843CrossRefPubMedPubMedCentralGoogle Scholar
  33. Kim J, Kim KP, Lim KT, Lee SC, Yoon J, Song G, Hwang SI, Schöler HR, Cantz T, Han DW (2015) Generation of integration-free induced hepatocyte-like cells from mouse fibroblasts. Sci Rep 5:15706CrossRefPubMedPubMedCentralGoogle Scholar
  34. Lalit PA, Salick MR, Nelson DO, Squirrell JM, Shafer CM, Patel NG, Saeed I, Schmuck EG, Markandeya YS, Wong R, Lea MR, Eliceiri KW, Hacker TA, Crone WC, Kyba M, Garry DJ, Stewart R, Thomson JA, Downs KM, Lyons GE, Kamp TJ (2016) Lineage reprogramming of fibroblasts into proliferative induced cardiac progenitor cells by defined factors. Cell Stem Cell 18:354–367CrossRefPubMedPubMedCentralGoogle Scholar
  35. Lee J, Sugiyama T, Liu Y, Wang J, Gu X, Lei J, Markmann JF, Miyazaki S, Miyazaki J, Szot GL, Bottino R, Kim SK (2013) Expansion and conversion of human pancreatic ductal cells into insulin-secreting endocrine cells. Elife 2:e00940PubMedPubMedCentralGoogle Scholar
  36. Lefebvre B, Belaich S, Longue J, Vandewalle B, Oberholzer J, Gmyr V, Pattou F, Kerr-Conte J (2010) 5’-AZA induces Ngn3 expression and endocrine differentiation in the PANC-1 human ductal cell line. Biochem Biophys Res Commun 391:305–309CrossRefPubMedGoogle Scholar
  37. Li H, Chen G (2016) In vivo reprogramming for CNS repair: regenerating neurons from endogenous glial cells. Neuron 91:728–738CrossRefPubMedGoogle Scholar
  38. Li W, Zhou H, Abujarour R, Zhu S, Young Joo J, Lin T, Hao E, Schöler HR, Hayek A, Ding S (2009) Generation of human-induced pluripotent stem cells in the absence of exogenous Sox2. Stem Cells 27:2992–3000PubMedPubMedCentralGoogle Scholar
  39. Li R, Liang J, Ni S, Zhou T, Qing X, Li H, He W, Chen J, Li F, Zhuang Q, Qin B, Xu J, Li W, Yang J, Gan Y, Qin D, Feng S, Song H, Yang D, Zhang B, Zeng L, Lai L, Esteban MA, Pei D (2010) A mesenchymal-to-epithelial transition initiates and is required for the nuclear reprogramming of mouse fibroblasts. Cell Stem Cell 7:51–63CrossRefPubMedGoogle Scholar
  40. Li Y, Zhang Q, Yin X, Yang W, Du Y, Hou P, Ge J, Liu C, Zhang W, Zhang X, Wu Y, Li H, Liu K, Wu C, Song Z, Zhao Y, Shi Y, Deng H (2011) Generation of iPSCs from mouse fibroblasts with a single gene, Oct4, and small molecules. Cell Res 21:196–204CrossRefPubMedGoogle Scholar
  41. Li H, Zhou H, Wang D, Qiu J, Zhou Y, Li X, Rosenfeld MG, Ding S, Fu X (2012) Versatile pathway-centric approach based on high- throughput sequencing to anticancer drug discovery. Proc Natl Acad Sci USA 109:4609–4614CrossRefPubMedPubMedCentralGoogle Scholar
  42. Li W, Cavelti-Weder C, Zhang Y, Clement K, Donovan S, Gonzalez G, Zhu J, Stemann M, Xu K, Hashimoto T, Yamada T, Nakanishi M, Zhang Y, Zeng S, Gifford D, Meissner A, Weir G, Zhou Q (2014a) Long-term persistence and development of induced pancreatic beta cells generated by lineage conversion of acinar cells. Nat Biotechnol 32:1223–1230CrossRefPubMedGoogle Scholar
  43. Li W, Nakanishi M, Zumsteg A, Shear M, Wright C, Melton DA, Zhou Q (2014b) In vivo reprogramming of pancreatic acinar cells to three islet endocrine subtypes. Elife 3:e01846PubMedPubMedCentralGoogle Scholar
  44. Li X, Zuo X, Jing J, Ma Y, Wang J, Liu D, Zhu J, Du X, Xiong L, Du Y, Xu J, Xiao X, Wang J, Chai Z, Zhao Y, Deng H (2015) Small-molecule-driven direct reprogramming of mouse fibroblasts into functional neurons. Cell Stem Cell 17:195–203CrossRefPubMedGoogle Scholar
  45. Lim KT, Lee SC, Gao Y, Kim KP, Song G, An SY, Adachi K, Jang YJ, Kim J, Oh KJ, Kwak TH, Hwang SI, You JS, Ko K, Koo SH, Sharma AD, Kim JH, Hui L, Cantz T, Schöler HR, Han DW (2016) Small molecules facilitate single factor-mediated hepatic reprogramming. Cell Rep 15:814–829CrossRefGoogle Scholar
  46. Lyssiotis CA, Foreman RK, Staerk J, Garcia M, Mathur D, Markoulaki S, Hanna J, Lairson LL, Charette BD, Bouchez LC, Bollong M, Kunick C, Brinker A, Cho CY, Schultz PG, Jaenisch R (2009) Reprogramming of murine fibroblasts to induced pluripotent stem cells with chemical complementation of Klf4. Proc Natl Acad Sci USA 106:8912–8917CrossRefPubMedPubMedCentralGoogle Scholar
  47. Minami K, Okuno M, Miyawaki K, Okumachi A, Ishizaki K, Oyama K, Kawaguchi M, Ishizuka N, Iwanaga T, Seino S (2005) Lineage tracing and characterization of insulin-secreting cells generated from adult pancreatic acinar cells. Proc Natl Acad Sci USA 102:15116–15121CrossRefPubMedPubMedCentralGoogle Scholar
  48. Muraoka N, Yamakawa H, Miyamoto K, Sadahiro T, Umei T, Isomi M, Nakashima H, Akiyama M, Wada R, Inagawa K, Nishiyama T, Kaneda R, Fukuda T, Takeda S, Tohyama S, Hashimoto H, Kawamura Y, Goshima N, Aeba R, Yamagishi H, Fukuda K, Ieda M (2014) MiR-133 promotes cardiac reprogramming by directly repressing Snai1 and silencing fibroblast signatures. EMBO J 33:1565–1581CrossRefPubMedPubMedCentralGoogle Scholar
  49. Nam YJ, Song K, Luo X, Daniel E, Lambeth K, West K, Hill JA, DiMaio JM, Baker LA, Bassel-Duby R, Olson EN (2013) Reprogramming of human fibroblasts toward a cardiac fate. Proc Natl Acad Sci USA 110:5588–5593CrossRefPubMedPubMedCentralGoogle Scholar
  50. Niu W, Zang T, Zou Y, Fang S, Smith DK, Bachoo R, Zhang CL (2013) In vivo reprogramming of astrocytes to neuroblasts in the adult brain. Nat Cell Biol 15:1164–1175CrossRefPubMedGoogle Scholar
  51. Pang Z, Yang N, Vierbuchen T, Ostermeier A, Fuentes DR, Yang T, Wernig M (2011) Induction of human neuronal cells by defined transcription factors. Nature 476:220–223PubMedPubMedCentralGoogle Scholar
  52. Pournasr B, Asghari-Vostikolaee MH, Baharvand H (2015) Transcription factor-mediated reprograming of fibroblasts to hepatocyte-like cells. Eur J Cell Biol 94:603–610CrossRefPubMedGoogle Scholar
  53. Qian L, Huang Y, Spencer CI, Foley A, Vedantham V, Liu L, Srivastava D (2012) In vivo reprogramming of murine cardiac fibroblasts into induced cardiomyocytes. Nature 485:593–598CrossRefPubMedPubMedCentralGoogle Scholar
  54. Rezvani M, Espanol-Suner R, Malato Y, Dumont L, Grimm AA, Kienle E, Willenbring H (2016) In vivo hepatic reprogramming of myofibroblasts with aav vectors as a therapeutic strategy for liver fibrosis. Cell Stem Cell 18:809–816CrossRefPubMedPubMedCentralGoogle Scholar
  55. Rie W, Muraoka N, Inagawa K, Yamakawa H, Miyamoto K, Sadahiro T, Umei T, Kaneda R, Suzuki T, Kamiya K, Tohyama S (2013) Induction of human cardiomyocyte-like cells from fibroblasts by defined factors. Proc Natl Acad Sci USA 110:12667–12672CrossRefGoogle Scholar
  56. Ring KL, Tong LM, Balestra ME, Javier R, Andrews-Zwilling Y, Li G, Walker D, Zhang WR, Kreitzer AC, Huang Y (2012) Direct reprogramming of mouse and human fibroblasts into multipotent neural stem cells with a single factor. Cell Stem Cell 11:100–109CrossRefPubMedPubMedCentralGoogle Scholar
  57. Sekiya S, Suzuki A (2011) Direct conversion of mouse fibroblasts to hepatocyte-like cells by defined factors. Nature 475:390–393CrossRefPubMedGoogle Scholar
  58. Shalem O, Sanjana NE, Hartenian E, Shi X, Scott DA, Mikkelsen TS, Heckl D, Ebert BL, Root DE, Doench JG, Zhang F (2014) Genome-scale CRISPR-Cas9 knockout screening in human cells. Science 343:84–87CrossRefPubMedGoogle Scholar
  59. Shi Y, Desponts C, Do JT, Hahm HS, Scholer HR, Ding S (2008) Induction of pluripotent stem cells from mouse embryonic fibroblasts by Oct4 and Klf4 with small-molecule compounds. Cell Stem Cell 3:568–574CrossRefPubMedGoogle Scholar
  60. Simeonov KP, Uppal H (2014) Direct reprogramming of human fibroblasts to hepatocyte-like cells by synthetic modified mRNAs. PLoS One 9:e100134CrossRefPubMedPubMedCentralGoogle Scholar
  61. Son EY, Ichida JK, Wainger BJ, Toma JS, Rafuse VF, Woolf CJ, Eggan K (2011) Conversion of mouse and human fibroblasts into functional spinal motor neurons. Cell Stem Cell 9:205–218CrossRefPubMedPubMedCentralGoogle Scholar
  62. Song K, Nam YJ, Luo X, Qi X, Tan W, Huang G, Acharya A, Smith CL, Tallquist MD, Neilson EG, Hill JA, Bassel-Duby R, Olson EN (2012) Heart repair by reprogramming non-myocytes with cardiac transcription factors. Nature 485:599–604CrossRefPubMedPubMedCentralGoogle Scholar
  63. Song G, Pacher M, Balakrishnan A, Yuan Q, Tsay HC, Yang D, Reetz J, Brandes S, Dai Z, Pützer BM, Araúzo-Bravo MJ, Steinemann D, Luedde T, Schwabe RF, Manns MP, Schöler HR, Schambach A, Cantz T, Ott M, Sharma AD (2016) Direct reprogramming of hepatic myofibroblasts into hepatocytes in vivo attenuates liver fibrosis. Cell Stem Cell 18:797–808CrossRefPubMedGoogle Scholar
  64. Tada M, Tada T, Lefebvre L, Barton SC, Surani MA (1997) Embryonic germ cells induce epigenetic reprogramming of somatic nucleus in hybrid cells. EMBO J 16:6510–6520CrossRefPubMedPubMedCentralGoogle Scholar
  65. Takahashi K, Yamanaka S (2006) Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell 126:663–676CrossRefPubMedGoogle Scholar
  66. Takahashi K, Yamanaka S (2016) A decade of transcription factor-mediated reprogramming to pluripotency. Nat Rev Mol Cell Biol 17:183–193CrossRefPubMedGoogle Scholar
  67. Vierbuchen T, Ostermeier A, Pang Z, Kokubu Y, Sudhof TC, Wernig M (2010) Direct conversion of fibroblasts to functional neurons by defined factors. Nature 463:1035–1041CrossRefPubMedPubMedCentralGoogle Scholar
  68. Wang H, Cao N, Spencer CI, Nie B, Ma T, Xu T, Zhang Y, Wang X, Srivastava D, Ding S (2014) Small molecules enable cardiac reprogramming of mouse fibroblasts with a single factor, Oct4. Cell Rep 6:951–960CrossRefPubMedPubMedCentralGoogle Scholar
  69. Wang Y, Qin J, Wang S, Zhang W, Duan J, Zhang J, Wang X, Yan F, Chang M, Liu X, Feng B, Liu J, Pei X (2016) Conversion of human gastric epithelial cells to multipotent endodermal progenitors using defined small molecules. Cell Stem Cell 19:449–461CrossRefPubMedGoogle Scholar
  70. Warren L, Manos PD, Ahfeldt T, Loh YH, Li H, Lau F, Ebina W, Mandal PK, Smith ZD, Meissner A, Daley GQ, Brack AS, Collins JJ, Cowan C, Schlaeger TM, Rossi DJ (2010) Highly efficient reprogramming to pluripotency and directed differentiation of human cells with synthetic modified mRNA. Cell Stem Cell 7:618–630CrossRefPubMedPubMedCentralGoogle Scholar
  71. Wen L, Tang F (2016) Single-cell sequencing in stem cell biology. Genome Biol 17:71CrossRefPubMedPubMedCentralGoogle Scholar
  72. Yamakawa H, Muraoka N, Miyamoto K, Sadahiro T, Isomi M, Haginiwa S, Kojima H, Umei T, Akiyama M, Kuishi Y, Kurokawa J, Furukawa T, Fukuda K, Ieda M (2015) Fibroblast growth factors and vascular endothelial growth factor promote cardiac reprogramming under defined conditions. Stem Cell Reports 5:1128–1142CrossRefPubMedPubMedCentralGoogle Scholar
  73. Yang H, Zhang L, An J, Zhang Q, Liu C, He B, Hao D (2016) MicroRNA-mediated reprogramming of somatic cells into neural stem cells or neurons. Mol Neurobiol. doi: 10.1007/s12035-016-0115-9 Google Scholar
  74. Yoo AS, Sun AX, Li L, Shcheglovitov A, Portmann T, Li Y, Lee-Messer C, Dolmetsch RE, Tsien RW, Crabtree GR (2011) MicroRNA-mediated conversion of human fibroblasts to neurons. Nature 476:228–231CrossRefPubMedPubMedCentralGoogle Scholar
  75. Yu B, He Z, You P, Han Q, Xiang D, Chen F, Wang M, Liu C, Lin X, Borjigin U, Zi X, Li J, Zhu H, Li W, Han C, Wangensteen KJ, Shi Y, Hui L, Wang X, Hu Y (2013) Reprogramming fibroblasts into bipotential hepatic stem cells by defined factors. Cell Stem Cell 13:328–340CrossRefPubMedGoogle Scholar
  76. Yuan X, Wan H, Zhao X, Zhu S, Zhou Q, Ding S (2011) Brief report: combined chemical treatment enables Oct4-induced reprogramming from mouse embryonic fibroblasts. Stem Cells 29:549–553CrossRefPubMedGoogle Scholar
  77. Yuan Y, Hartland K, Boskovic Z, Wang Y, Walpita D, Lysy PA, Zhong C, Young DW, Kim YK, Tolliday NJ, Sokal EM, Schreiber SL, Wagner BK (2013) A small-molecule inducer of PDX1 expression identified by high-throughput screening. Chem Biol 20:1513–1522CrossRefPubMedPubMedCentralGoogle Scholar
  78. Zhang L, Yin J, Yeh H, Ma N, Lee G, Chen X, Wang Y, Lin L, Chen L, Jin P, Wu G, Chen G (2015) Small molecules efficiently reprogram human astroglial cells into functional neurons. Cell Stem Cell 17:735–747CrossRefPubMedPubMedCentralGoogle Scholar
  79. Zhang M, Lin Y, Sun Y, Zhu S, Zheng J, Liu K, Cao N, Li K, Huang Y, Ding S (2016a) Pharmacological reprogramming of fibroblasts into neural stem cells by signaling-directed transcriptional activation. Cell Stem Cell 18:653–667CrossRefPubMedPubMedCentralGoogle Scholar
  80. Zhang Y, Cao N, Huang Y, Spencer CI, Fu J, Yu C, Liu K, Nie B, Xu T, Li K, Xu S, Bruneau BG, Srivastava D, Ding S (2016b) Expandable cardiovascular progenitor cells reprogrammed from fibroblasts. Cell Stem Cell 18:368–381CrossRefPubMedGoogle Scholar
  81. Zhao Y, Londono P, Cao Y, Sharpe EJ, Proenza C, O’Rourke R, Jones KL, Jeong MY, Walker LA, Buttrick PM, McKinsey TA, Song K (2015a) High-efficiency reprogramming of fibroblasts into cardiomyocytes requires suppression of pro-fibrotic signalling. Nat Commun 6:8243CrossRefPubMedPubMedCentralGoogle Scholar
  82. Zhao Y, Zhao T, Guan J, Zhang X, Fu Y, Ye J, Zhu J, Meng G, Ge J, Yang S, Cheng L, Du Y, Zhao C, Wang T, Su L, Yang W, Deng H (2015b) A XEN-like state bridges somatic cells to pluripotency during chemical reprogramming. Cell 163:1678–1691CrossRefPubMedGoogle Scholar
  83. Zhou W, Freed CR (2009) Adenoviral gene delivery can reprogram human fibroblasts to induced pluripotent stem cells. Stem Cells 27:2667–2674CrossRefPubMedGoogle Scholar
  84. Zhou Q, Tripathi P (2012) How to remake a fibroblast into a neural stem cell. Cell Stem Cell 10:347–348CrossRefPubMedGoogle Scholar
  85. Zhou Q, Brown J, Kanarek A, Rajagopal J, Melton DA (2008) In vivo reprogramming of adult pancreatic exocrine cells to beta-cells. Nature 455:627–632CrossRefPubMedGoogle Scholar
  86. Zhou Y, Wang L, Vaseghi HR, Liu Z, Lu R, Alimohamadi S, Yin C, Fu J, Wang G, Liu J, Qian L (2016) Bmi1 is a key epigenetic barrier to direct cardiac reprogramming. Cell Stem Cell 18:382–395CrossRefPubMedPubMedCentralGoogle Scholar
  87. Zhu S, Li W, Zhou H, Wei W, Ambasudhan R, Lin T, Kim J, Zhang K, Ding S (2010) Reprogramming of human primary somatic cells by OCT4 and chemical compounds. Cell Stem Cell 7:651–655CrossRefPubMedGoogle Scholar
  88. Zhu S, Ambasudhan R, Sun W, Kim HJ, Talantova M, Wang X, Zhang M, Zhang Y, Laurent T, Parker J, Kim HS, Zaremba JD, Saleem S, Sanz-Blasco S, Masliah E, McKercher SR, Cho YS, Lipton SA, Kim J, Ding S (2014a) Small molecules enable OCT4-mediated direct reprogramming into expandable human neural stem cells. Cell Res 24:126–129CrossRefPubMedGoogle Scholar
  89. Zhu S, Rezvani M, Harbell J, Mattis AN, Wolfe AR, Benet LZ, Willenbring H, Ding S (2014b) Mouse liver repopulation with hepatocytes generated from human fibroblasts. Nature 508:93–97CrossRefPubMedPubMedCentralGoogle Scholar
  90. Zhu S, Wang H, Ding S (2015) Reprogramming fibroblasts toward cardiomyocytes, neural stem cells and hepatocytes by cell activation and signaling-directed lineage conversion. Nat Protoc 10:959–973CrossRefPubMedGoogle Scholar
  91. Zhu S, Russ HA, Wang X, Zhang M, Ma T, Xu T, Tang S, Hebrok M, Ding S (2016) Human pancreatic beta-like cells converted from fibroblasts. Nat Commun 7:10080CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© The Author(s) 2017

Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made.

Authors and Affiliations

  1. 1.Life Sciences InstituteZhejiang UniversityHangzhouChina

Personalised recommendations