Ever since their Nobel Prize-winning discovery by Rita Levi-Montalcini and Stanley Cohen in the 1950s [5,6], growth factors have been reported to play increasingly diverse and important roles in cell growth and differentiation. These roles lend them importance not only in development but also in regeneration after injury [3] (clinicaltrials.org: NCT00000842). Epidermal growth factor (EGF) is found to enhance healing of wounds in the skin and cornea [7]. Nerve growth factor (NGF) and brain-derived or glial cell-derived neurotropic factors (BDNF and GDNF, respectively) have been shown in rat models of nerve damage to enhance neuronal regeneration [8,9]. Similarly, platelet-rich plasma (PRP) has been shown in numerous animal studies to enhance tissue regeneration after injuries [10]. The plasma extracted from blood is enriched in the laboratory for platelets, which contain presynthesized growth factors packaged in α-granules. Upon activation of clotting (after infusion into the injured tissue) these growth factors are secreted into the tissue microenvironment. Autologous transplant of PRP for the treatment of various degenerative diseases of tissue or bone has reached clinical trials (clinicaltrials.gov: NCT00761423, NCT01355549). Although the results from many of the trials and existing clinical reports have been promising (but inconsistent or ambiguous at best [11,12]), there still seems to be anticipation that with proper standardization of the techniques for plasma isolation, enrichment, storage, and delivery, better and more consistent outcomes can be expected. Pro-angiogenic factors such as vascular–endothelial growth factor (VEGF), fibroblast growth factor (FGF), and platelet-derived growth factor (PDGF) are another subclass of growth factors that have been utilized to improve regeneration by inducing neovascularization at the site of injury [13,14].

The immune system plays an important role in the resolution of injury. Early after injury, it clears the site of injury of debris and infectious agents, and allows repair to begin. However, persistent and uncontrolled inflammation can be detrimental to wound repair [15], so drugs that modulate the inflammatory response have long been utilized to enhance it. Thrombin peptide (THP508) was reported to enhance fracture healing through activation of tumor necrosis factor-α (TNFα) and other pro-inflammatory factors [16]. On the other hand, attenuation of inflammatory response has also been shown to enhance regenerative repair. Several reports have shown that administration of high doses of nonsteroidal anti-inflammatory drugs improve regeneration of brain tissue after the ischemic injury induced by a stroke in rodents [17,18]. Even the ability of stem-cell therapy using mesenchymal stromal or stem cells (MSCs) to enhance tissue regeneration is attributed in part to their immune-modulatory properties [19].

The extracellular matrix (ECM) plays a key role in each step of injury repair. Within days after injury, a fibrous matrix is synthesized to replace lost or damaged tissue. The matrix is remodeled over time as the repair progresses. All the steps, from inflammation to reepithelialization to contraction, involve very close interactions with matricellular proteins, which sequester growth factors, regulate collagen turnover, and provide scaffold and support to allow cells to grow during tissue repair [20]. ECM components, such as heparin, bind many signaling molecules and growth factors and enhance their signaling by sequestering them to the microenvironment or by stabilizing them. They have been used to enhance signaling by proteins of interest [21]. Moreover, a polymer mimetic of heparan sulfate proteoglycan itself has demonstrated remarkable healing properties in both topical wounds and internal injuries [22,23]. A clinically available formulation is already in use in Europe and the Middle East to treat corneal ulcers and diabetic wounds [24]. Since heparan sulfates are important for the signaling of growth factors and cytokines, their degradation and cleavage in a wound microenvironment impairs healing. The heparan sulfate mimetics are uncleavable by heparanases and glycanases and hence aid repair of the wounded tissue [25].

A small molecule is a low-molecular-weight (smaller than 500 Daltons) organic compound. Small molecules that are investigated for use as therapeutics typically bind to a protein, nucleic acid, or polysaccharide and alter its enzymatic activity or biological function [26]. Small molecules are identified by cell phenotypic, reporter-based, or organism-based screening of chemical libraries [27]. Phenotypic screening is based on the cellular phenotype or cell-surface markers in response to the chemical library. In order to identify the modulators of specific pathways, promoter-driven reporters are used as readouts for the pathways of interest. Zebrafish and xenopus are routinely used for organism-based screens [28]. There is increasing interest in finding small molecules that can modulate signaling pathways as they have a number of advantages over biologics such as proteins and antibodies. For one thing, they are cheaper to synthesize. Due to their small size, they are often orally bioavailable, and hence delivery may not require injection. Additionally, they are more likely to be cell-permeable and thus to allow modification of the pathway at multiple steps; an antibody would rely solely on the sequesterization of a ligand or the blocking of the receptor. Hence, small molecules are very useful in blocking intracellular protein–protein interactions, which has to date been the sole domain of antisense oligos. Small molecules thus offer a safer, more effective, and easier alternative to traditional signaling pathway-modulation approaches [26].

Given their superiority in modifying molecular signals, many types of small molecule have been investigated for potential roles in regeneration. Small-molecule antioxidants such as quercetin, allopurinol retinoids, and uric acid are reported to enhance wound healing [29]. Ascorbic acid, another small molecule, commonly used for stem-cell differentiation, is also known to regulate ECM by increasing collagen synthesis [30]. In a model of combined injury (fracture/irradiation) in mice, Greenberger and colleagues showed that the small molecule GS-Nitroxide attenuates ionizing radiation-driven delay in fracture healing [31]. Another small molecule, Pirfenidone, has been evaluated in clinical trials as an antifibrotic agent for idiopathic pulmonary fibrosis. Already reported to reduce fibrosis in lung, liver, heart, and kidney through the modulation of the transforming growth factor beta (TGFβ) pathway, the molecule has potential for therapeutic application in tissue injury, through promoting regeneration by minimizing scarring [32]. Work by our group has also shown that in a mouse myocardial infarct model, treatment with the small-molecule Wnt inhibitor Pyrvinium alleviates adverse cardiac remodeling post-infarct [33]. The same molecule, upon topical treatment on mouse-ear punch injury, caused dramatic closure of the holes with remarkable restoration of tissue structure, including hair-follicle regeneration [34]. However, careful and thorough screenings of these molecules need to be performed before they can be proposed for therapeutic purposes, since some may have toxicity and off-target effects. Nonetheless, with careful considerations of their shortcomings, the exciting prospects for small molecules in advancing regenerative medicine cannot be overlooked.

As the field of regenerative medicine has become more multidisciplinary, innovative approaches to drug delivery, in vitro organ regeneration, and cell therapy have been introduced. These new technologies have revolutionized the field of regenerative medicine by increasing the efficacy and versatility of the traditional methods of enhancing regeneration through cell- or drug-based approaches.

Tissue engineering has been an important part of the field of regenerative medicine for many decades now. With the design and application of sophisticated biomaterials, tissue engineering is moving even further toward improving regenerative therapy [35]. Degradable biomaterials composed of collagen, fibronectin, chitosan, and so on have been used for the delivery of cells and to provide ECM components for the survival or directed differentiation of injected cells. Alginate, an algae-derived polysaccharide, has been used in the form of sponge or gel to enhance the regeneration of peripheral nerve [36], spinal cord [37], or excised axons [38] in rodent or feline models when used alone or in combination with neural progenitor (neurosphere) cells [39]. In a remarkable study, Atala and colleagues used a cell-polymer matrix composed of a Poly (Lactide-co-Glycolide)-poly(D,L-lactide-co-glycolide) (PGA-PLGA) scaffold lined with urothelial cells on one side and muscle cells on the other to show for the first time the reconstitution of an autonomous and functional hollow organ using a tissue-engineering approach [40].

Another avenue in which bionics and biomaterial science are contributing to better tissue regeneration is in the sustained delivery of stabilized growth factors or drugs. In 2003, Hyongbum Kim et al. reported successfully using PLGA scaffolds for the slow release over a month of ascorbate-2-phosphate (AsAP) and dexamethasone to induce osteogenesis from MSCs [41]. Another recent study showed that an injectable polyvalent coacervate of a polycation and heparin used as a matrix for the sustained delivery and stabilization of FGF2 increased FGF2-mediated angiogenesis in a rodent model [21].

In order to be useful for clinical purposes, stem cells should maintain their self-renewal ability and be able to differentiate into specific lineages. Several biologics and genetic manipulations have been used to control the fate of stem cells, driving their self-renewal, differentiation, survival, or reprogramming. However, modulation of these complex biological events with small molecules seems to be safer and more effective in regulating stem-cell fate. Using high-throughput screening of chemical libraries, many molecules have been identified that target specific cellular pathways to maintain stem-cell self-renewal, direct stem-cell differentiation, and enable or enhance stem-cell reprogramming [27, 52]. Here we will focus on recent interventions introduced by small molecules in affecting and/or improving the behavior of different types of stem cells commonly used in regenerative biology.