Cell dedifferentiation is the process by which cells grow reversely from a partially or terminally differentiated stage to a less differentiated stage within their own lineage. This extraordinary phenomenon, observed in many physiological processes, inspires the possibility of developing new therapeutic approaches to regenerate damaged tissue and organs. Meanwhile, studies also indicate that dedifferentiation can cause pathological changes. In this review, we compile the literature describing recent advances in research on dedifferentiation, with an emphasis on tissue-specific findings, cellular mechanisms, and potential therapeutic applications from an engineering perspective. A critical understanding of such knowledge may provide fresh insights for designing new therapeutic strategies for regenerative medicine based on the principle of cell dedifferentiation. Earthworms readily reform two new duplicates after being cut in half2. Some amphibians, such as newts, create new limbs, tails and jaws with identical structures or functions to the lost body parts after amputation3, which is regarded as complete regeneration. However, at the organismal level, humans  have a limited capacity for regeneration. The structure or functions of a damaged organ may be repaired to a compromised extent compared to the native status, and such “incomplete” regeneration usually occurs within this specific organ4. Typical cases include scarring in the skin that restores coverage without perspiration, as well as fibrocartilage that provides a structure different from articulate cartilage (and therefore inferior functions). Thus, these complete regenerative phenomena are prompting researchers to determine the underlying mechanisms and transform this mythological image into reality for humans. Dedifferentiation is regarded as one of the mechanisms involved in regeneration, as it enables cells, especially those without proliferative potential, to proliferate again and redifferentiate, leading to the replacement of the lost cells.Regeneration has always been a dream of humans, as shown in various fairy tales. Some invertebrates and amphibians have extraordinary capacity for regeneration. Zebrafish are able to fully regenerate their hearts, even after 20% of the ventricle is removed5. During differentiation, the fundamental basis of development of an organism, a less specialized cell type gradually transforms into a more specialized cell type, which is constrained to a stable morphology, structure and function. Moreover, the process of differentiation reduces the self-renewal ability and pluripotency of the cell. Differentiation continues during adulthood to maintain homeostasis. Dedifferentiation is a cellular process by which cells grow in reverse, from a partially or terminally differentiated stage to a less differentiated stage within their own lineage. In general, the phenomenon is manifested by a change in the shape, gene expression pattern, protein expression pattern and function. For example, dedifferentiated chondrocytes experience a change in phenotype after repeated culture in monolayers. The phenotype of spherical chondrocytes changes to spindle-shaped fibroblastic-like cells. Meanwhile, dedifferentiation results in a shift in the expression of the type II collagen (Col II) gene to type I collagen (Col I) and thus a corresponding change in the production of Col II protein to Col I protein in the extracellular matrix (ECM)6. In addition, these changes also lead to a switch from hyaline cartilage to fibrous cartilage, which would considerably change the function of articular cartilage7.There are a variety of cell types throughout multicellular organisms. All of these cells originate from a simple zygote through a series of processes such as cell division and differentiation8. Terminally differentiated cells obtained in a noninvasive manner are potentially transformed to pluripotent stem cells through dedifferentiation. Then, these cells would be propagated in vitro, differentiated into objective cells and transplanted in vivo to achieve the regeneration of the damaged organ. Based on accumulating evidence, signaling pathways play a critical role in the process of dedifferentiation. Activation of the Wnt/β-catenin signaling pathway induces the dedifferentiation of epidermal cells9, articular chondrocytes10, or endothelial cells11 for regeneration. In addition, some specific biomolecules trigger the dedifferentiation of vascular smooth muscle cells12 or chondrocytes13 via the mitogen-activated protein kinase (MAPK) pathways. Also, innovative biomaterials are designing and creating to regulate dedifferentiation15. As our understanding of the molecular mechanisms of dedifferentiation improves, therapeutic methods using dedifferentiation present tremendous potential.In recent decades, an increasing number of researchers have proposed that the induction of dedifferentiation shows promise to repair injured tissue in the clinicIn this review, we compile the literature on recent advances in research of dedifferentiation, with an emphasis on the current knowledge of dedifferentiation processes, possible mechanisms and the therapeutic applications with an engineering perspective. A comprehensive understanding of such knowledge may provide fresh insights into the potential use of dedifferentiation in translational science.16. Here, the dedifferentiation of mammalian cardiomyocytes and neurons, which have been studied extensively, will be discussed in detail.The phenomena of dedifferentiation exists in various tissues and organs from plants, amphibians and animals, manifesting mainly as re-entry into the cell cycle, the acquisition of a stem cell-like phenotype, the expression of stem cell markers and redifferentiation to regenerate damaged tissues17. However, Porrello et al.18 observed a transient regenerative potential of the neonatal murine heart induced by the dedifferentiation of cardiomyocytes. Moreover, cardiomyocyte remodeling, which includes the restructuring of the normally existing structures and rearrangement of the cytoskeleton, is vital for the normal function of heart19. A recently proposed hypothesis states that cardiomyocyte remodeling is closely related to dedifferentiation, which is characterized by a switch in the expression pattern of mature cardiomyocytes to a more immature state, enabling cardiomyocytes to survive under unfavorable conditions, enter the cell cycle to proliferate and recover their functions21. MacLellan’s group demonstrated in mouse model that dedifferentiation may be an essential prerequisite for cardiomyocyte proliferation22. Both early genes, such as α-smooth muscle actin (α-SMA) and GATA4, and characteristic markers that are generally expressed in stem cells, such as Runx1 and Dab2, are re-expressed in dedifferentiated cardiomyocytes, which might facilitate proliferation and improve the multipotent differentiation capacity23. The dedifferentiation of cardiomyocytes may be an alternative pathway to induce regeneration, as observed in the newt, whose heart is regenerated through the dedifferentiation of mature cardiomyocytes and subsequent proliferation , a member of the interleukin-6 family, in the mechanism regulating cardiomyocyte dedifferentiation. OSM triggers the Ras/MEK/Erk cascade to induce dedifferentiation of in vitro rat cardiomyocyte via OSM receptor β. A positive feedback loop mediates murine cardiomyocyte dedifferentiation; it comprises Yes-associated protein (YAP), transcriptional enhancer factor (TEAD1) and OSM—where the YAP-TEAD1 pathway stimulates OSM expression, OSM activates YAP-TEAD1, and OSM also upregulates OSM receptors through the YAP-TEAD1 pathway26. The size of OSM-treated cardiomyocytes also increases, which may allow them to re-establish cellular contacts to promote cellular survival27.An in vitro model in which adult rabbit cardiomyocytes were cocultured with cardiac fibroblasts was established28. The findings from numerous studies on the role of RB are consistent with this finding. The inactivation of RB, which is attributed to phosphorylation by increased numbers of cyclin D1/cdk4 complexes, is related to cardiomyocyte hypertrophy in congestive heart failure29. According to MacLellan et al.30, mice lacking RB and RB-like 2 have a larger heart and exhibit increased cardiomyocytes proliferation. Zaglia et al.31 were the first researchers to report that GATA4, a zinc-finger transcription factor, is expressed by fibroblasts in the adult rat heart, but not in fibroblasts from the gut, skin and skeletal muscle. In addition, the authors established an in vitro coculture experiment with cardiac fibroblasts and cardiomyocytes. Cardiac fibroblasts enhance the dedifferentiation of adult cardiomyocytes, a change that is strongly correlated with cell cycle reprogramming, as evidenced by the expression of DNA synthesis markers. Stimulation of GATA4 by overexpression of GRP78 (glucose-regulated protein of 78 kDa) can promote cardiomyocyte growth in mice32. However, further studies are needed to investigate the underlying mechanisms by which GATA4 affects cell division.In addition, during the process of limb regeneration in urodele amphibians, the retinoblastoma (RB) protein has a very important role in facilitating the re-entry of differentiated cells into the cell cycle. During this process, mature cells dedifferentiate through the inactivation of RB and then enter the cell cycle33. Schwann cells remove myelin and axonal debris after dedifferentiation by themselves or by recruiting circulating macrophages34. Upon dedifferentiation, Schwann cells downregulate myelin-associated genes that are essential for the myelination process, such as myelin protein zero and Krox20/Egr-235, and re-express molecules associated with immature states, such as p75 neurotrophin receptor (p75NTR), neural cell adhesion molecule (NCAM) and L136. Moreover, Schwann cells produce neurotrophic factors to support axon regeneration37  pathways in Schwann cell dedifferentiation. Although extracellular signal-regulated kinase (ERK) signaling is implicated in Schwann cell dedifferentiation39, increased ERK phosphorylation alone is not sufficient to cause dedifferentiation without colony-stimulating factor (CSF)-1-activated macrophage reactions40. The transcription factor c-Jun, which is the terminal component of the Jun-N-terminal kinase (JNK) pathway, is a negative regulator of myelination41. Parkinson et al.42 genetically depleted c-Jun from murine Schwann cells and observed a delay in demyelination after injury and the suppression of dedifferentiation. Recently, the transient reactivation of mTOR complex 1 was shown to be necessary to increase c-Jun translation and Schwann cell dedifferentiation after mice nerve injury43. Additionally, c-Jun contributes to the production of neurotrophic factors, including glial cell line-derived neurotrophic factor (GDNF) and Artemin, which is beneficial for axonal regeneration45. The activation of p38 MAPK induces c-Jun expression and drives rat Schwann cell dedifferentiation46. Park and colleagues47 reported an important role for the Rac1 GTPase (Rac) in Schwann cell dedifferentiation. Rac controls the demyelination of murine Schwann cells in Schmidt-Lantermann incisures and induces c-Jun expression in injured sciatic nerves. Rac also regulates the expression of GDNF and p75NTR, which are related to regeneration. Furthermore, the neuregulin-Rac-MKK7-JNK/c-Jun pathway is required for Schwann cell dedifferentiation following rat nerve injury48. As shown in the study by Jung’s group, extracellular adenosine triphosphate inhibits rat Schwann cell proliferation and Schwann cell dedifferentiation by blocking the lysosomal activation49. Likewise, increased cAMP levels induce and maintain the differentiation of Schwann cells, while the removal of cAMP rapidly induces the dedifferentiation and proliferation of Schwann cells50. Moreover, according to Viader et al.51, miRNAs expressed in Schwann cells play a key role in modulating murine Schwann cell response to nerve injury. The authors identified interactions of miR-34a with Notch1 and Ccnd1 to promote the dedifferentiation and proliferation of Schwann cells, and miR-140 influenced the remyelination.Based on accumulating evidence, several intracellular signaling pathways are involved in Schwann cell dedifferentiation. Notch signaling negatively regulates myelination, accelerates the dedifferentiation of mature Schwann cells and promotes the proliferation of immature Schwann cells53. Conditioned medium from scratch-insulted astrocytes (ACM) induces astrocytes to regain NSPCs potentials and these NSPCs of astrocytic origin redifferentiated into neuronal and glial cells55. Overexpression of Ezh2 increases the expression of neural stem cell genes and downregulates the expression of astrocytic genes, but these changes were insufficient to induce a complete dedifferentiation56. A series of studies has been conducted by Yang et al.57 to investigate the underlying mechanisms involved in the astrocyte dedifferentiation process. Using an in vitro scratch-wound model, the authors revealed that ACM treatment stimulated the rat astrocytes to dedifferentiate into radial glial progenitor cells with the characteristic radial glial-like morphology, re-expression of nestin, RC2 and paired homeobox proteins, and increased multipotent differentiation capability. Concomitantly, the progressive increase in the level of ErbB2 protein level suggested that the activation of the ErbB2 signaling pathways may lead to all these phenotypes in astrocytes. Furthermore, fibroblast growth factor 4 (FGF4), which is actively involved in neurogenesis58, is required for the astrocyte dedifferentiation because it activates the PI3K/Akt/p21 signaling pathway59. Astrocytes lacking FGF4 do not undergo dedifferentiation, while the application of FGF4 alone to astrocyte cultures fails to elicit the reprogramming of astrocytes towards NSPCs. In addition, in vitro experiments confirmed that the increase in sonic hedgehog (Shh) production after invasive injury plays a key role in rat astrocyte dedifferentiation60. The authors observed a significant inhibition of astrocyte dedifferentiation into NSPCs using a neutralizing antibody to inhibit Shh. Moreover, the addition of Shh alone had little effect on the dedifferentiation process and reversion to NSPCs. Based on these results, Shh is necessary but not sufficient to induce astrocyte dedifferentiation. However, in a subsequent study, Sirko drew a distinct conclusion that Shh signaling is necessary and sufficient to promote the proliferation of astrocytes in a mouse model and trigger their ability to form neurospheres in vitro61. More recent studies further demonstrate that Shh signaling effectively promotes conversion of murine astrocytes to a pluripotent state63. The treatment with two chemically distinct agonists of the Shh signaling promoted the elongation and proliferation of murine astrocytes64. The mechanism was related to the crosstalk between Sox2/Shh-targeted downstream signals and PI3K/Cdk2/Smurf2 signaling65.In addition, astrocytes, which are widely distributed throughout the central nervous system (CNS), dedifferentiate into neural stem/progenitor cells (NSPCs) upon exposure to appropriate stimuli based on compelling evidenceAs mentioned above, the induction of dedifferentiation might be beneficial for tissue recovery in various organs. However, it might also cause some pathological events in certain tissues. In this case, cartilage is one of the classic tissue models and will be discussed in details.66. Chondrocyte dedifferentiation induces substantial changes in the levels of cell surface markers and cytoskeletal proteins, as well as the appearance of nonspecific ECM, which would ultimately lead to osteoarthritis. In addition to changes in the aggrecan to versican and Col II to Col I ratios and the deposition of collagen type X67, which are markers of the chondrocyte dedifferentiation status, the ratio of the cell surface markers CD14 to CD90 at the mRNA and protein levels has been corroborated as a new cell membrane-based differentiation index68. In addition, CD166 is an indicator of chondrocyte redifferentiation69. As shown in the study by Hong and Reddi, microRNA-221 and microRNA-222 are upregulated during dedifferentiation, while microRNA-140, microRNA-143 and microRNA-145 are downregulated70. Moreover, chondrocyte dedifferentiation controls the mechanical properties of the cell, such as increasing cell stiffness by enhancing membrane-actin adhesion71. However, chondrocyte dedifferentiation is not related to cell division72.Chondrocyte dedifferentiation has some similarities to osteoarthritis, such as a significant decrease in the expressions of aggrecan and Col II73 investigated the role of MAPK signaling in the dedifferentiation of in vitro primary bovine chondrocyte using inhibitors to block ERK, JNK, and p38. The inhibition of ERK and JNK by their respective inhibitors PD98059 and SP600125 significantly increased the expression of both chondrogenic marker genes and fibrotic genes, while blockade of p38 with SB203580 significantly upregulated Col II expression but inhibited Col I expression. Thus, p38 has a substantial contribution to the dedifferentiation of primary chondrocytes under normal monolayer culture conditions. Interestingly, Kim and colleagues74 reported that cytokine-induced apoptosis inhibitor-1 (CIAPIN-1) induced the dedifferentiation of in vitro cultured rabbit articular chondrocytes by activating ERK-1/2 and inactivating p38. The inhibition of ERK-1/2 by PD98059 appears to suppress the CIAPIN-1-induced dedifferentiation, while the addition of SB203580 to inhibit p38 promotes dedifferentiation. This inconsistency is possibly because—(i) other signaling molecules activated and (ii) the extent of p38 inhibition. The latter could be more likely because this group further observed that p38 inhibition under both CIAPIN-1 promoter and inhibitor SB203580 enhanced dedifferentiation. Two more subsequent findings by this group76 are similar, both involving a pre-inducing factor. Kim and colleagues also performed a series of in vitro studies in this area. They investigated the effects of thymoquinone (TQ) and PEP-1-SIRT2 on the dedifferentiation and inflammation of rabbit chondrocytes77. TQ or PEP-1-SIRT2 induce the expression of cyclooxygenase-2 (COX-2), causing the dedifferentiation of rabbit articular chondrocytes78 and subsequently leading to the dedifferentiation of chondrocytes through the ERK pathways. TQ-induced inflammation is mediated by the phosphoinositide 3-kinase (PI3K) and p38 pathways, while PEP-1-SIRT2-induced inflammation is mediated by the p38 and ERK pathways. The authors also reported that salinomycin (SAL) causes the dedifferentiation of rabbit articular chondrocytes via the ERK pathway, as determined by increased ERK activity without alterations in JNK and p38 activity following treatment with SAL79. Recently, the authors revealed that PEP-1-glutaredoxin-1 increased endoplasmic reticulum-stress and then stimulated the dedifferentiation of rabbit chondrocytes via the ERK-1/2 pathway75. Another Korean lab clarified the molecular mechanisms by which interleukin (IL)-1β, a proinflammatory cytokine associated with various cellular activities, induced the dedifferentiation of in vitro cultured primary rabbit chondrocytes80. IL-1βinduces the expression of nicotinamide phosphoribosyltransferase (NAMPT) and then activates SIRT1, which subsequently induces the activation of ERK and p38. In turn, ERK and p38 activate SIRT1, forming a positive feedback loop81. Integrin αvβ5 was reported to activate ERK signaling and to subsequently enhance the dedifferentiation of in vitro cultured primary human articular chondrocytes82. In addition, the Notch pathway is involved in the regulation of matrix metalloproteinase 13 (MMP-13) activity and the differentiation of human articular chondrocytes in vitro83. In rabbit articular chondrocytes in vitro, 2-deoxy-D-glucose (2DG) induces dedifferentiation via the β-catenin pathway84. A recent study revealed a feed-forward loop driven by USP14 (a deubiquitinating enzyme) and the NF-kappa B pathway that enhances the effect of IL-1β on the dedifferentiation of human chondrocytes in vitro85.An increasing number of studies have been conducted to explore the cytokines and signaling pathways that are involved in chondrocyte dedifferentiation. MAPK signaling, consisting of ERK, JNK and p38, has very important roles in chondrocyte phenotype and metabolism. Rosenzweig et al.86. The disruption of the actin cytoskeleton inhibits the dedifferentiation of in vitro cultured rabbit chondrocytes by blocking PKCα and –ζ signaling87. The integrity of the actin cytoskeleton is related to the differentiated phenotype of chondrocytes through the PI3K/Akt and p38 pathways88. In addition, the Rho family GTPase ROCK plays a key role in organizing the actin cytoskeleton89. Treatment with a ROCK inhibitor alters human chondrocyte morphology in vitro and activates SOX9 and chondrogenic gene expression, suggesting the suppression of dedifferentiation90. Berberine, a compound widely present in many plants, reorganizes the actin cytoskeleton and subsequently induces dedifferentiation via PI3K/Akt and p38 pathways91. Focal adhesion kinase (FAK) is involved in cell adhesion, migration and cell proliferation. The role of FAK in the chondrocytic dedifferentiation process was investigated by Shin et al.92 An increasing number of passages in monolayer culture resulted in higher levels of focal adhesion complexes. In addition, FAK knockdown facilitates the restoration of the expression of cartilage-specific genes, including Col II, aggrecan and SOX9, in rat dedifferentiated chondrocytes in vitro. In another study, FAK knockdown inhibited porcine chondrocyte proliferation in vitro, and FAK was required to modulate the expression of Col II93. Ding and colleagues94 further explored the effects of spreading areas and aspect ratios of single chondrocytes undergoing dedifferentiation using a micropatterning technique in vitro. Rat chondrocytes incubated under conditions of larger sizes and higher aspect ratios preferentially undergo dedifferentiation.The architecture of the actin cytoskeleton modulates the phenotype of chondrocytes during dedifferentiation16. Meanwhile, dedifferentiated chondrocytes gradually lose the ability of proliferation and undergo hypertrophy and eventually apoptosis95. Therefore, understanding the impact of cell dedifferentiation in specific tissue is essential for devising further therapeutic strategies—such as induction versus suppression (and to what extent). To date, much remains unknown about the causes and consequences of dedifferentiation under specific physiological or pathological circumstances, which requires more mechanistic studies at different levels.The information above provides interesting implications. First, dedifferentiation may have either positive or negative consequences to tissue repair in different tissue/organs. For example, after injury, dedifferentiated cardiomyocytes were preferentially protected from apoptosis; they can survive and re-enter cell cycle for tissue repair8. Inspired by the studies on dedifferentiation of amphibians such as newt, researchers found that murine myotubes could dedifferentiate to a certain extent when stimulated by an extract obtained from newt regenerating limbs96. Also, partial dedifferentiation of Schwann cells can be induced by local factors such as hormones and neurotrophic factors97. These results suggest that such extrinsic signals can play an important role in dedifferentiation. Although current interpretations remain on a rather speculative level and await further verification, in vitro investigations on dedifferentiation provide valuable insights into the molecular mechanisms and the pathways associated with cell dedifferentiation, with extensive in vivo studies in demand.Second, it should be noted that the findings from non-human models provide valuable evidence for assessing the application potential of dedifferentiation for future trials and translations in humans. Many studies show that a main reason for the huge difference of regenerative capacity between amphibians and mammals is the induction of dedifferentiation, which does not often occur after injury in mammals98. Dedifferentiation refers to a reversion of a fully differentiated cell within its own lineage, which allows the cell to proliferate again to replace those damaged cells in many cases99. Trans-differentiation, first reported in the newt for regeneration of lens over 100 years ago, involves dedifferentiation in the first step and subsequent differentiation into another cell type. It may also be observed during experimental induction of trans-differentiation that cells can directly trans-differentiate to a distinct lineage through an “unnatural” intermediate phase. Thus, a strategy for regenerative medicine would be either (i) to induce dedifferentiation and then proliferation of target cells or (ii) to stimulate trans-differentiation of cells into the desired cell types. In either case, a clear dissection of the changes and impacts of the genes involved in dedifferentiation is fundamental.Third, by exploring the regenerative phenomena in invertebrates and amphibians, researchers have uncovered the cellular and molecular mechanisms to develop regenerative strategies for humans. Dedifferentiation and trans-differentiation, part of the physiological response to injury in numerous organs, are mechanistically associated with natural regeneration100, immunoregulatory function101, and multilineage differentiation potential102, MSC-based therapies for tissue regeneration have not yet been accepted clinically103. Current efforts focus on the development of bioreactor system, which could combine scaffolds, biochemical factors, and mechanical stimuli to simulate the in vivo microenvironment so that MSCs-based cellular products can meet the requirements for clinical use104. Recent studies have provided evidence that dedifferentiation is a promising approach for tissue regeneration4. An increasing number of trials have used engineering approaches to regulate dedifferentiation.Although mesenchymal stem cells (MSCs) are promising “seed” cells in tissue regeneration owing to their excellent characteristics such as self-renewal capacity105. Recently, researchers have focused on designing and preparing innovative biomaterials to induce dedifferentiation. Yahalom-Ronen et al.15 established an in vitro culture system for murine P1 neonatal cardiomyocytes. They constructed softer substrates to stimulate dedifferentiation and the cell cycle re-entry of cardiomyocytes, suggesting that the development of soft scaffolds might facilitate the regeneration of the adult human heart. A keratin hydrogel was created and used to repair a 1-cm defect in rat sciatic nerve injury models106. Compared with Matrigel, the keratin hydrogel group induced more rapid dedifferentiation of Schwann cells, leading to the faster activation of macrophages and more efficient clearance of myelin debris and ECM remodeling. Thus, the positive regulation of Schwann cell dedifferentiation encourages effective nerve regeneration.Scaffold materials, which can provide a three-dimensional culture condition that more closely resembles the in vivo environment, play a critical role in the biological characteristics and proliferation rate of seeding cells, subsequently determining the fate and function of cells107 developed a “continuous expansion” (CE) in vitro culture technique that avoids repeated passaging and maintains the chondrocytic phenotype. Using this new method, bovine chondrocytes were cultured on high-extension silicone rubber (HESR) dishes with continuously expanding and elastic properties. Furthermore, the authors functionalized the HESR dishes with a cartilage ECM extract from a cow108. HESR dishes functionalized with a cartilage extract were more effective at inhibiting chondrocytic dedifferentiation than control dishes. Another group further developed biomimetic materials using decellularized ECM derived from human articular chondrocytes14. These biomaterials were more effective at maintaining the phenotype and enhancing the proliferation of chondrocytes than standard culture plates. Moreover, similar results were obtained using MSC-derived ECM110, indicating that a cell type-specific ECM is more similar to the native microenvironment and reduces the dedifferentiation of chondrocytes. Gelatin methacryloyl/hyaluronic acid methacrylate hydrogels displayed their positive effects on down-regulation of dedifferentiation gene markers111. In addition, a biomimetic hydrogel was prepared based on alginate and tuned by altering the concentration of chlorosulfonic acid112. The sulfation of alginate, which mimics proteoglycans in cartilage, promotes in vitro bovine chondrocyte proliferation, increase cartilage matrix deposition, and inhibits the expression of dedifferentiation markers. A blended gel composed of alginate and collagen also provided a preferable gel matrix environment for the suppression of rat chondrocyte dedifferentiation in vitro113. More interestingly, Ding and colleagues114 concentrated on the effects of external nanoscale features on the dedifferentiation of in vitro cultured primary rat chondrocytes. Poly(ethylene glycol) (PEG) hydrogels were modified with nanopatterned arginine–glycine–aspartate (RGD) and used to culture chondrocytes  direct addition in the form of recombinant proteins and (ii) gene delivery of their complementary DNAs (cDNAs) using viral or nonviral vectors.115. Based on these findings, the group subsequently performed in vivo experiments; GM6001 was injected into rats immediately after sciatic nerve crush to stimulate Schwann cell division, reduce myelin protein expression and subsequently support axonal regeneration116. In another interesting study, leukemia inhibitory factor (LIF), a promyelination factor, was encapsulated in nanoparticle carriers and delivered to enhance remyelination and promote nerve regeneration117, highlighting the feasibility of biochemically regulating tissue development with engineering approaches. Similarly, recombinant human midkine was added to the growth medium of rat chondrocytes for in vitro monolayer expansion and not only promoted cell proliferation but also suppressed dedifferentiation, avoiding a shift in the chondrocyte to fibroblast phenotype and decreasing the expression of chondrocytic genes118. In addition, andrographolide, the main active component of the traditional Chinese medicine Andrographis paniculata, also prevents the dedifferentiation of rabbit articular chondrocytes in vitro, and 3 μM was reported to be the optimal dose119. Although positive effects have been observed by directly supplementing culture media with biomolecules, this method will result in instability in vivo, limiting further clinical applications. Therefore, gene-transfer strategies appear to be a promising alternative.MMPs, which are capable of degrading ECM proteins, regulate Schwann cell signaling and mitosis. As shown in a study by the Shubayev group, MMP-9 inhibits Schwann cell mitosis during nerve regeneration, while the MMP inhibitor, GM6001, counteracts the effect of MMP-9 and further increases the division of murine Schwann cells120. Therefore, researchers have constructed adenoviral vectors to express a truncated mutant form of eNO synthase followed by an intraneural injection of the adenovirus121. Rat Schwann cell dedifferentiation was induced, as evidenced by the overexpression of dedifferentiated Schwann cell marker growth-associated protein 43 and an increase in the bands of Bungner, the cellular substrate for guiding growing axons. On the other hand, various biomolecules are being developed to inhibit the dedifferentiation of chondrocytes. Based on the results of the previous studies showing that IL-1β induces the dedifferentiation of chondrocytes and is inhibited by cytokine response modifier A (CrmA)123, hyaluronic acid-chitosan microspheres were designed for the controlled release of CrmA124. Rat chondrocytes treated with these microspheres in vitro maintain the chondrocytic phenotype and production of cartilage-specific proteins even after cocultured with IL-1β. Additionally, chondrocytes expressing wild-type p53-inducible phosphatase (Wip1) following microporation generally retain a chondrocyte phenotype for a longer time during in vitro expansion125.Two main types of gene delivery vectors have bee identified: viral vectors, such as adenoviral vectors and lentiviral vectors, and nonviral vectors, such as polymers and liposomes. Viral vectors have better transfection efficiencies while nonviral vectors have advantages, such as ease of synthesis and low immunogenicity. Endothelial nitric oxide (eNO) plays a pivotal role in regulating Schwann cell dedifferentiation. NO synthesized by eNO synthase decelerates axon regeneration after crush of the XII nerve126 to modulate the expression of a constitutively active (ca) form of ErbB2 in murine cardiomyocytes following the administration of the tetracycline analog doxycycline (DOX). Notably, caErbB2 promotes the dedifferentiation and subsequently induces cell division, highlighting the proliferative and regenerative potential of adult cardiomyocytes.A “Tet-Off” transgenic mouse model was established by Tzahor and colleagues127 explored the therapeutic potential of the dedifferentiated MSCs (De-MSCs) in neuron repair  to human chondrocytes in vitro, resulting in an increase in the DNA methylation level of the Col I-A1 promoter, which slowed chondrocyte dedifferentiationDiscovering the interesting phenomenon of cell dedifferentiation, followed by uncovering its underlying mechanisms, opens new avenues for controlling cell fate for regenerative applications. It is possible that in different tissue/organs, dedifferentiation occurs in various frequencies and to different extents; and perhaps much remains unknown about the causes and consequences of this phenomenon under specific physiological or pathological circumstances. However, there must be a reason that the organisms develop the capability of dedifferentiation through the long period of evolution—to adapt, to survive, to heal, to evade from dangers, and to evolve. As such, recapitulating dedifferentiation in regenerative medicine may provide a new, rational, evolution-inspired strategy to bypass inherent limitations with therapeutic cells (such as inability to proliferate) and initiate tissue development under better control.Future studies can be directed in three aspects. First, more mechanistic studies at different levels are required to understand the initiation of cell dedifferentiation. Genetic, epigenetic, transcriptional, metabolic, and mechanobiological controls of the cell fate decision towards dedifferentiation are all worth investigation, which will offer comprehensive clues for designing engineering tools. Second, tissue regeneration involves multiple cell types of different lineages and characteristics, especially in the case of a complex organ such as the liver, kidney, and heart. It is very likely that the change in the behavior of one cell type affects the function of another. Hence, the consequence of engineered dedifferentiation of a group of target cells on other cells in the tissue niche should be carefully examined. Any potential adverse effects—either short-term  or long-term  risks—need to be avoided. Third, for any practice in which cell dedifferentiation demonstrates preliminary signs of safety and effectiveness, extensive work is demanded to standardize the protocol. Major questions include: how far should the cells be reversed along the way of dedifferentiation, how many percentages of the cells are effectively induced to dedifferentiate, and when is the best timing to apply the dedifferentiated cells into tissue repair. As such, exciting future work lies ahead from biological, clinical, and engineering perspectives for translating the discovery of cell dedifferentiation into tissue regeneration.


The impact load, such as seismic and shock wave, sometimes causes severe damage to the reinforced concrete structures. This study utilized different lengths of chopped carbon fibers to develop a carbon-fiber-reinforced mortar (CFRM) and carbon-fiber-reinforced concrete (CFRC) with high impact and anti-shockwave resistance. The different lengths  of chopped carbon fibers were pneumatically dispersed and uniformly mixed into the cement with a 1% weight proportion. Then the CFRM and CFRC specimens were made for static and dynamic tests. The compressive and flexural strengths of the specimens were determined by using the standard ASTM C39/C 39M and ASTM C 293-02, respectively. Meanwhile, a free-fall impact test was done according to ACI 544.2R-89, which was used to test the impact resistances of the specimens under different impact energies. The CFRM and CFRC with a length of 6 mm exhibit maximum compressive strength. Both flexural and free-fall impact test results show that the 24 mm CFRM and CFRC enhances their maximum flexural strength and impact numbers more than the other lengths of CFRM, CFRC, and the benchmark specimens. After impact tests, the failure specimens were observed in a high-resolution optical microscope, to identify whether the failure mode is slippage or rupture of the carbon fiber. Finally, a blast wave explosion test was conducted to verify that the blast wave resistance of the 24 mm CFRC specimen was better than the 12 mm CFRC and benchmark specimens. Reinforced concrete structures are sometimes subjected to seismic and high impact loadings, which might cause catastrophic damage. An impact load might be an aircraft taking off or landing on a runway, or a heavy vehicle passing over a bridge expansion joint; in a harbor, a ship may collide with a wharf due to the movement of the waves. These impact loadings cause damage, such as cracks and spalling, to the reinforced concrete pavement. Generally, natural and manmade fibers incorporated into reinforced concrete structures can improve the durability and toughness and can reduce the shrinkage of concrete. Recently, carbon-fiber-reinforced mortar (CFRM) and carbon-fiber-reinforced concrete (CFRC) are usually being used in the repair, rehabilitation, and rebuilding work of civil engineering infrastructures.Fiber-reinforced mortar (FRM) enhances the tensile and impact resistance, and fiber is used as reinforcement to increase its strength. Concrete is a brittle material and prone to cracks or damage when subjected to an external force, and the load-bearing capacity and its serviceability will be reduced. If the compression and impact resistance of concrete structures can be improved, then the service life of the various reinforced concrete structures can be prolonged. Fiber-reinforced concrete (FRC) has good mechanical properties and is used in multiple construction environments.In the last few decades, many studies have been conducted on the performance of different fibers applied in mortar and concrete. Concrete reinforced by steel rebars and polypropylene fiber has shown an increase in mechanical performance on flexural strength and improved ability to impact resistance . Fiber-rSeveral studies reveal the effect of adding different fibers on the mechanical properties of fiber-reinforced concrete: The impact resistance of self-consolidating rubberized concrete (SCRC) can be determined by the reinforced steel fiber and synthetic semi-rigid fiber; the addition of synthetic semi-rigid fiber to self-filling rubber concrete can improve impact resistance. Geopolymer concrete reinforced with mono fiber has a remarkable enhancement in its impact strength and fracture toughness. It was concluded that the steel fiber–polypropylene fiber reinforced specimen has the best toughness, with a proportion of 1.5%, and it takes repeated impacts to reach complete failure ,9,10,11.Carbon fiber is a lightweight composite material, which does not corrode, degrade, or fatigue. Due to its high specific strength, it is used in the aerospace industry, for automotive parts and sports equipment, and in civil engineering. The content and length of carbon fiber in concrete affect performance on the strength of various aspects. As the amount and length of additive carbon fiber increases, the impact resistance significantly improves. The failure of fiber-reinforced concrete paste is mainly due to the force endured beyond the bonded force between surfaces ,15,16,17Chopped carbon fiber was used in this study, to determine the static (compression and bending) and impact behavior of concrete and mortar with different lengths of fiber . The carbon fibers were separated by dispersion and then incorporated into the cement with a 1% weight proportion. A pneumatic process was additionally used to disperse the carbon fiber for uniform distribution in the concrete and mortar.The carbon fibers were chopped at different lengths , which were uniformly mixed in slurry forms for mortar and concrete. In this section, the materials, experimental methods, and equipment are listed; they include various material properties of carbon fiber, aggregates, and cement. Carbon-fiber tensile strength and elastic modulus are superior to other fibers; the properties are listed in Portland cement provides overall strength, and it is used within one month after the manufacture date. Note that the cement should not be in direct contact with the ground, in order to avoid moisture absorption. The carbon fiber is included in a cement matrix, in a dry state composition.Carbon fiber applications are exploited in the aerospace industry, automobile industry, and other fields, which have high specific strength and fatigue resistance. The short carbon fiber was acquired from Tairylan Division, Formosa Plastics Group, Taipei, Taiwan, R.O.C. , and theThe carbon fiber surface microscopy was examined with a scanning electron microscope , at the Department of Molecular Science and Engineer Lab, National Taipei University of Technology. The carbon-fiber-surface SEM observation is shown in Fiber-reinforced mortar (FRM) presents high flexural deformation; it can increase durability, reduce shrinkage, and improve toughness. The carbon fiber can restrain mortar cracking and prevent fracture failure; it can also offer improvement to the mechanical and physical properties of mortar. In this study, the water–cement ratio of 0.4 was used in the CFRM specimens, and sand was 105% of the cement weight. The dispersed carbon fiber was mixed in the dry stage and then wet state, to aid in the uniform distribution in the mortar.Generally, fibers are usually used in concrete, to control cracking due to plastic shrinkage and drying shrinkage. Some types of fibers could produce greater impact resistance, abrasion resistance, and shatter resistance in concrete. Since the modulus of elasticity of carbon fiber is higher than it is for the plain concrete, it can help the CFRC carry the load by increasing the tensile strength. To ensure that each fiber strand is effective, it is necessary to disperse the fibers uniformly in the concrete. In this study, the water–cement ratio of the CFRC was 0.45, and the sand weight was the same as the CFRM. As the component of a composite material that resists compressive stress, the aggregate was 225% of the cement weight. The fineness modulus of aggregates for CFRC specimen was 6.01, as shown in A series of tests were conducted according to ASTM and ACI standards ,33,34, t−6/s to 10−4/s), which was acted on the flat surface of the specimen, and the dimension was ⌀10 cm × 20 cm [The carbon-fiber-reinforced composites were tested under ASTM C39/C 39M-01 standards . The cyl × 20 cm , respectAccording to ASTM C 293-02 , the CFRThe free-fall impact tests of CFRM and CFRC were conducted under ACI 544-2R . The sieThe CFRM and CFRC were tested with different impact energies; the string was used to hang the mass steel ball at a certain height. The CFRM and CFRC specimens’ surfaces resist a single impact at high energy and repeated impact at lower energy. The impact energy can be represented as follows.E is potential energy (J), m is mass (kg), g is gravity acceleration (m/s2), and h is height (m). The CFRC and CFRM are examined at different energies with a single and repeated free-fall impact test. The CFRC and CFRM with different lengths of fiber  possess better mechanical performance than the benchmark.In Equation (1), The disparate lengths of fiber 6, 12, and 24 mm were incorporated with cement, to prepare CFRM and CFRC specimens. The test results of compressive, bending, and impact performance were obtained with the different fiber lengths of carbon fiber in CFRM and CFRC specimens.In this subsection, the CFRM and CFRC compressive strengths were tested with different lengths of fiber and then compared with a benchmark specimen (without added carbon fiber). Each test group had three specimens.The strength of the CFRM is higher than that of the benchmark mortar specimen, and increasing compressive strength is dependent on the length of the carbon fiber. The chopped 6 mm carbon fibers were uniformly inhabited in the cylindrical specimens because the length of 6 mm fibers is shorter than the others, and it occupies more volume than the longer fiber in the concrete matrix with the same weight proportion. The compressive strength of CFRM specimen was increased by reducing the length of the carbon fiber.In The C-L6-C chopped carbon-fiber-reinforced concrete has a maximum compressive strength than the benchmark and other fiber-reinforced concrete (such as C-L24-C and C-L12-C). Because the length of 6 mm fibers is shorter than the others, they possess more volume than the longer fibers in the concrete matrix with the same weight proportion; that is, 6 mm fiber occupies two times more space than the 12 mm carbon fibers, and four times more space than the 24 mm carbon fibers.The chopped CFRM and CFRC increase their flexural strength, compared to the standard benchmark specimens. In this subsection, the flexural strength of CFRM and CFRC with different lengths of fiber are compared with the benchmark specimens (without added carbon fiber). The flexural strength of fiber-reinforced mortar and concrete is discussed below. Each test group has three specimens.As shown in From the above test results, we can see that the length of the long carbon fiber (24 mm) enhances its maximum strength in the CFRM. This is because the longer carbon fiber can resist tensile force, and its failure mode is fracture failure instead of slip failure , with a high magnification range between 200 and 400, at the material laboratory of the Institute of Mineral Resources Engineering, National Taipei University of Technology. The CFRC fracture surface from the photomicrograph is obtained from an optical microscope, as shown in The blast wave explosion test was used to verify the anti-blast wave resistance of different carbon fiber lengths of CFRC. In Among them, the C-L6-C specimen exhibits good compressive strength, which is 25.3% (40.28 Ma) more than the C-B-C specimen. The C-L12-C and C-L24-C specimens also increase the compressive strength by 14.4% and 4.4%, respectively. The inclusion of 6 mm carbon fiber exhibits the maximum compressive strength. Compared to the C-B-M specimen, the C-L6-M, C-L12-M, and C-L24-M specimens enhance their compressive strength by 22.2%, 14%, and 11.3%, respectively. The compressive-strength-enhancement effect decreases by increasing the length of the fiber.The flexural strength of the F-L24-M specimen increases its strength, compared with the benchmark, by 42.14%. The CFRM specimens F-L6-M and F-L12-M also enhance their strength by 24.57% and 29.06% respectively. The CFRC specimens F-L24-C improve flexural strength by 27.07 MPa. The specimens F-L6-C (17.82%) and F-L12-C (22.92%) also increase their strength, compared with the benchmark (F-B-C), which shows that flexural strength increases as the length of the chopped carbon fibers increases.The impact number of the CFRM specimen exhibits that the specimen I-L24-M can sustain high impact energy at 339 J, and specimen I-L24-M has maximum impact numbers, as compared with specimens I-L12-M and I-L6-M, at different impact energies. The I-L24-C specimen exhibits higher impact-energy resistance, as compared with the benchmark and other specimens, at different impact energies. The failure impact numbers of the CFRM and CFRC specimens show that the strength is increased by increasing the chopped-carbon-fiber length.In the blast test, the specimen BW-B-C exhibits crushing failure, and the specimens BW-L12-C and BW-L24-C exhibit spalling failure. The spalling depth of specimen BW-L12-C is 5 cm, and the inner/outer circle diameters of the spalling surfaces are 36 and 50 cm, respectively. Similarly, the spalling depth of specimen BW-L24-C is 4.3 cm, and the inner/outer circle diameters of the spalling surfaces are 31 and 35 cm. The test result indicates that the specimen BW-L24-C has better anti-blast performance than other specimens.The CFRC test results show that CFRC can be applied in the reinforced concrete structures of bridge expansion joints and aircraft runways, due to its excellent compressive, bending, and impact performances. The CFRC can be applied in the reinforced-concrete structures, to prevent damage from the blast wave and seismic loading.The CFRM test results show that CFRM can be applied in the repair work in the reinforced concrete structure, due to its excellent mechanical performance.The chopped carbon fibers  enhanced the static and impact load performances of mortar and concrete. The compressive strength increased with a decrease in the length of chopped carbon fibers. The pneumatic dispersion process also aids in distributing the fiber uniformly. The 24 mm chopped carbon-fiber-reinforced cement composites better resist repeated impacts at low energy than the composites reinforced by other lengths of carbon fibers. The conclusions are as follows:


The gut microbiome (GMB) generates numerous small chemicals that can be absorbed by the host and variously biotransformed, incorporated, or excreted. The resulting metabolome can provide information about the state of the GMB, of the host, and of their relationship. Exploiting this information in the service of biomarker development is contingent on knowing the GMB-sensitivity of the individual chemicals comprising the metabolome. In this regard, human studies have lagged far behind animal studies. Accordingly, we tested the hypothesis that serum levels of chemicals unequivocally demonstrated to be GMB-sensitive in rodent models would also be affected in a clinical patient sample treated with broad spectrum antibiotics.We collected serum samples from 20 hospitalized patients before, during, and after treatment with broad-spectrum antibiotics. We also collected samples from 5 control patients admitted to the hospital but not prescribed antibiotics. We submitted the samples for a non-targeted metabolomic analysis and then focused on chemicals known to be affected both by germ-free status and by antibiotic treatment in the mouse and/or rat.Putative identification was obtained for 499 chemicals in human serum. An aggregate analysis did not show any time x treatment interactions. However, our literature search identified 10 serum chemicals affected both by germ-free status and antibiotic treatment in the mouse or rat. Six of those chemicals were measured in our patient samples and additionally met criteria for inclusion in a focused analysis. Serum levels of 5 chemicals  declined significantly in our group of antibiotic-treated patients but did not change in our patient control group.Clostridial species. We conclude that judiciously selected serum chemicals can reliably detect antibiotic-induced suppression of the GMB in man and thus facilitate further metabolome-based biomarker development.Broad-spectrum antibiotic treatment in patients lowered serum levels of selected chemicals previously demonstrated to be GMB-sensitive in rodent models. Interestingly, all those chemicals are known to be uremic solutes that can be derived from aromatic amino acids  by anaerobic bacteria, particularly  The gut microbiome (GMB) generates numerous small  chemicals that can be absorbed, biotransformed, and either incorporated or excreted by the host –3. The lDevelopment of such potential biomarkers requires identifying the GMB-sensitive constituents of the metabolome. While germ-free mice and rats (GF) have proved invaluable in this regard , they haClostridium difficile and vancomycin-resistant Enterococci  AND metabolome.” We evaluated the resulting articles as well as any additional relevant publications that they referenced. Antibiotic reports were considered to be relevant if they met the following criteria, i) they used a conventional animal strain, ii) the antibiotic was administered for > 6 days, iii) a control group not treated with antibiotic was included, iv) animals were maintained on a standard lab diet, and v) blood was drawn no later than 24 hours after completion of antibiotic treatment. Our final focused list consisted of chemicals whose serum levels were reported to be affected both by GF status and by treatment with at least 1 antibiotic.a priori to exclude those chemicals for which > 50% of baseline data were missing. Given that antibiotics often suppress GMB-derived metabolites, chemicals with > 50% missing values at the post-antibiotic time point were included. To address fluctuations that can occur for reasons other than antibiotic treatment, we also decided a priori to exclude those chemicals which showed significant changes between any of the time points, PRE-, MID-, and POST-antibiotic in the control group. Finally, chemicals that were significantly different between the control group and antibiotic group at the PRE time point were also to be excluded a priori.From the data provided by Metabolon©, we decided a priori exclusions. Chemicals common to both lists were subjected to planned t-test comparisons with significance set at P < 0.05, uncorrected for multiple comparisons.The list of chemicals obtained from the PubMed search was cross-referenced with the list of chemicals we measured in patient serum subject to the A total of 20 antibiotic-treated patients  and 5 controls  completed the study. With the exception of 1 antibiotic-treated participant who provided 2 MID blood samples and no POST sample, all other participants provided PRE, MID, and POST samples. The antibiotic treatment regimens included a broad-spectrum beta-lactam antibiotic  alone in 4 patients, and in combination with additional agents in 16 patients (14 intravenous vancomycin and 2 moxifloxacin). The median duration of antibiotic treatment was 5 days (range 2-20 days).The metabolomic analysis provided putative identification for 499 chemicals. Under general analysis, 69 chemicals in the control group and 153 chemicals in the antibiotic group showed significant uncorrected POST vs PRE differences , but in Our PubMed search identified 8 relevant studies of antibiotic-effects on serum chemical levels in mice or rats –50 Tabl. We alsoEight such chemicals were among those measured in our human serum . N-acetyIn our group of patients treated with broad-spectrum antibiotics, serum levels of 5 of 6 chemicals predicted to be highly GMB-dependent by animal studies, declined significantly as measured by univariate analysis. While consistent with our hypothesis, the findings have to be critically interpreted in terms of assumptions, experimental limitations, and generalizability.We enlisted inpatient male veterans with any of several medical conditions for which broad-spectrum antibiotic treatment was indicated. The duration of antibiotic treatment varied considerably. Our small sample of controls was not clinically matched with the treatment group. We used a commercial metabolomic service (Metabolon©) that conducted a non-targeted analysis using GC/MS or LC/MS/MS. While such an approach supports detection of a very large number of chemicals, it does so on the basis of available spectroscopic libraries rather than by direct comparisons to authentic standards. This yields presumptive identification and semi-quantitative comparisons . On the P < 0.05 accepted, relatively few analytes would have reached significance. This can be partially attributed to the high variance of the data as well as to the unequal sizes of the control (n = 5) and antibiotic-treated (n = 20) groups. We dealt with these issues by imposing criteria determined a priori to restrict the number of chemicals admitted to a focused univariate analysis.A classical ANOVA that included all the aggregate data  did not show any time x treatment interactions that exceeded the preset q < 0.1. Part of this was due to the FDR for multiple corrections. However, even if the q value was ignored and any time x treatment interaction a priori to exclude chemicals whose levels in the control group showed significant changes during the experimental period. It turned out, however, that this rule had no practical effect and did not further limit the chemicals identified as antibiotic-responsive in animal studies (Table 2). Second, we decided to exclude chemicals that at the PRE  time point already tended to be significantly different (P < 0.1) between controls and antibiotic-treated individuals. Such differences may be attributable to chance, underlying illness, or other factors. L-tyrosine was excluded in this way. Third, we decided to exclude all chemicals for which > 50% of data were missing for any time point in the controls or at the PRE time point in the antibiotic group. This was based on the fact that GF-status ) yielded only 5 heterogeneous human serum studies of antibiotic effects –17. An 8There are many steps between the appearance of a molecule in the gut lumen, whether as an endogenous or microbial product, and its entry into peripheral serum . TranspoThe serum chemicals we identified as antibiotic-sensitive in man belong to the phenolic  or indolic  pathways. Hippuric acid is formed through the conjugation of benzoic acid (BA) with the hydrophilic moiety glycine , 77 in aWhile HA and its hydroxylated metabolites are constituents of the human fecal metabolome , the GMBSimilarly, neither phenol nor p-cresol can be synthesized in appreciable quantities by endogenous human enzymes  but can While antibiotic treatment is not known to affect conjugation of BA or p-cresol; effects on their transport must be considered separately. Gut luminal BA crosses the basilar membrane of enterocytes via monocarboxylic transporters , 112. AnSulfated conjugates of phenol and p-cresol can be detected in human feces , 116. InConstituents of the human GMB readily convert L-TRP to indole , 132–134The kinetics of indole propionic acid are less well characterized. While human cells can successively metabolize L-TRP to indole-3-pyruvic acid and then to indole-3-lactic acid , 143, on5HT was the only chemical whose serum levels were affected both by GF and antibiotic status in rodents (Table 2) but which was not significantly affected by antibiotic treatment in our focused analysis (Table 3). It is also the only chemical in our list that can be generated endogenously by mammalian cells . IsolateShort chain fatty acids (SCFAs), generated via GMB-mediated fermentation of dietary fiber and protein, upregulate tryptophan hydroxylase 1 , 155. Sia priori criteria for selecting chemicals of interest we did not include data from patients with ileostomies. While the overwhelming preponderance of the total GMB reside in the colon, the bacterial density in the terminal ileum actually exceeds that of the large intestine [In developing our ntestine . Furtherntestine , 161. Nantestine . The enzntestine . At leasntestine . This suntestine  of 3 of ntestine , 166.Without any direct measures of the GMB, we cannot empirically evaluate possible associations between antibiotic-induced changes in the serum metabolome and concomitant changes in individual operational taxonomic units (OTUs) of the GMB. The available literature supports several generalizations. Most antibiotic-treatment regimens lower the overall diversity of the GMB , 167–170per se carry the fingerprint of the particular microbe that generated it. Hence, antibiotic-induced lowering of serum chemicals is more likely to be detected for those chemicals generated relatively exclusively by taxa targeted by the antibiotic regimen.Most metabolic pathways are encoded within functional genetic categories distributed across multiple types of bacteria rather than being exclusive to one genus or strain , 67, 171Bacteroides, Lactobacillus, Clostridia, and Bifido-bacteria [Clostridium difficile can ferment L-TYR, L-PHE, or L-TRP whereas C. perfringens can do so only to L-TYR [C. difficile is the only Clostridial species expressing 4-hydroxyphenylacetate decarboxylase (EC 4.1.1.83), an enzyme critical to the metabolism of L-TYR through to the end product p-cresol [All of the 5 chemicals determined to be antibiotic-responsive in our study are themselves either fermentation products or metabolites of fermentation products of aromatic amino acids. Although aromatic amino acids can be metabolized to some degree by cultures of a range of intestinal anaerobes, including members of the genera bacteria , only libacteria , 98–102.to L-TYR , 144. Fup-cresol , 174.E. coli, express tryptophanase (EC 4.1.99.1), which converts L-TRP to indole [Clostridium difficile cannot itself generate indole but can modify the intestinal milieu to promote indole-generation by other bacteria [Clostridial strains, can ferment L-TRP to indole propionic acid [A relatively large number of anaerobic constituents of the healthy human GMB, including e EC 4.1..1, whicho indole , 132–134bacteria . A more e EC 4.1..1, whichnic acid , 143–147Clostridial species [We noted earlier, that many small phenolic molecules can also be generated by GMB-mediated degradation of polyphenol rich dietary components –89. This species –179. Reg species , dietary species , 182. A  species . This unin vitro monoculture experiments must be complemented by in vivo studies that allow for polymicrobial population dynamics. Such efforts remain highly preliminary. In a subsample (n = 855) of individuals with minimal renal decline, serum indoxyl sulfate and p-cresol sulfate were both associated with the presence of family members of Clostridiales in the GMB [Clostridiales [Clostridia family.Of course, data from  the GMB . More reridiales . Thus, oClostridia and other species. All 5 chemicals, often referred to as uremic solutes were previously considered to be GMB-dependent based on various animal and human studies [Using a clinical sample of patients and a limited number of targets we demonstrated that broad-spectrum antibiotic treatment lowered serum levels of 5 chemicals  previously shown to be absent or markedly suppressed in the serum of GF mice (Table 3). Based on a consideration of transport and other mechanisms, we conclude that the most likely explanation for the data is an antibiotic-induced suppression of  studies , 166. Ho studies , socioec studies , genetic studies , concomi studies , and par studies  and colo studies . It is o studies . The faca priori could allow detection of effects that would not be apparent if statistical adjustments for comparing a very large number of variables were applied. Rapidly emerging data implicating the microbiome and its dependent metabolome across medical disciplines, including immunology [In view of the increasing use of antibiotics worldwide , metabolmunology  and neurmunology , oblige This work was supported by the Research Service of the VA Northeast Ohio Healthcare System and by a grant from the VISN 10 Research Initiative Program (to CJD and GEJ). The manuscript content is solely the responsibility of the authors and does not necessarily represent the official views of the Department of Veterans Affairs and/or the authors' employers and/or their academically affiliated institutions.


Background: The novel coronavirus disease 2019 (COVID-19) has impacted many countries across all inhabited continents, and is now considered a global pandemic, due to its high rate of infectivity. Research related to this disease is pivotal for assessing pathogenic characteristics and formulating therapeutic strategies. The aim of this paper is to explore the activity and trends of COVID-19 research since its outbreak in December 2019.Methods: We explored the PubMed database and the World Health Organization (WHO) database for publications pertaining to COVID-19 since December 2019 up until March 18, 2020. Only relevant observational and interventional studies were included in our study. Data on COVID-19 incidence were extracted from the WHO situation reports. Research output was assessed with respect to gross domestic product (GDP) and population of each country.Results: Only 564 publications met our inclusion criteria. These articles came from 39 different countries, constituting 24% of all affected countries. China produced the greatest number of publications with 377 publications (67%). With respect to continental research activity, Asian countries had the highest research activity with 434 original publications (77%). In terms of publications per million persons (PPMPs), Singapore had the highest number of publications with 1.069 PPMPs. In terms of publications per billion-dollar GDP, Mauritius ranked first with 0.075.Conclusion: COVID-19 is a major disease that has impacted international public health on a global level. Observational studies and therapeutic trials pertaining to COVID-19 are essential for assessing pathogenic characteristics and developing novel treatment options. The capital of the Chinese province Hebei, Wuhan city, has witnessed starting the 31st of December 2019, the emergence of a new lower respiratory tract disease .Between January and March 2020, worldwide efforts have been focused on dealing with this emerging pandemic . The higThat mentioned, research done on COVID-19 is of major importance for both the containment of the disease and the treatment of the patients . ReportsThe PubMed database of the National Center for Biotechnology Information (NCBI) and WHO database were used to find the publications related to this study. In PubMed, publications were identified by searching for the terms “novel coronavirus 2019,” “coronavirus 2019,” “COVID 2019,” and “COVID 19” in the search field. All publications between December 2019 and March 16, 2020 were included.Data were also extracted from the WHO database of publications on COVID-19, which is collected from bibliographic databases, table of contents of relevant journals, and other relevant scientific articles. PubMed publications were cross-referenced to the ones found in the WHO spreadsheet on COVID-19 publications. All the relevant articles not present on PubMed were then added to our database. News reports that were present in the WHO database were identified by two reviewers, and were removed from our final database.Four reviewers (from the authors) looked at the publications. The corresponding author’s country of origin was identified. The publication type was identified and only original articles and case reports were included in our study. The types of studies included basic science studies, epidemiological studies, randomized control trials, prospective trials, retrospective studies, and case series and reports. Descriptive analysis was done to report the number and type of articles from each country. The number of articles in each country was then compared to its number of confirmed cases to identify countries where more publications are needed.Data from the WHO Health Emergency Dashboard novel COVID-19 situation reports were extracted, from its initiation  up until the most recent one  . This inOur search yielded 1809 publications between December 16, 2019 and March 16, 2020. After removing duplicates, news articles, and articles not related to the topic as deemed by two independent reviewers, we selected 1596 publications for our analysis. Of these, only 564 publications (35%) met our inclusion criteria , Singapore ranked first with 1.069 PPMPs, followed by Mauritius with 0.791 PPMPs (Table Number of studiesCOVID-19 has arguably been the main focus of medical and scientific institutions around the globe for the past few months, and that is evident by the release of 1596 publications pertaining to it since December 16, 2019. Nevertheless, only around 35% of those constitute observational and interventional studies. China alone has contributed over 66% of the 564 publications, followed by the United States with 39 publications, or almost 7% of the total. For a topic of the utmost medical importance, we may expect to have a larger number of original publications; including but not limited to randomized trials and case series. With the rapid spread of COVID-19 around the globe, healthcare systems and experts were required to mount a rapid response as well. As clinical interventions and contingency planning rose in priority , the timOver the course of history, humanity has faced a plethora of deadly diseases. From smallpox and the bubonic plague, to AIDS and Ebola, the human race is no stranger to global pandemics -18. As mCountries with abundance of casesThe countries with the largest number of cases are China, Italy, Iran, Spain, and Germany in descending order. Considering the fact that China houses over 3.61 million licensed physicians, and is the birthplace of the current pandemic , it is nOf the current 54 countries with 100 or more reported COVID-19 cases, we notice that only five countries have published 10 or more articles. With limited knowledge of this novel virus, the value of case-series and observational studies in countries with a large number of cases should not be underestimated; hence, medical institutions should be contacted and encouraged to publish their findings. Population-level data can be a valuable tool for designing medical management algorithms and guidelines.Value of observational and interventional studiesAs history can attest to, humanity has been surviving epidemics with improved outcomes. This is largely due to our most important line of defense, information. The quicker quality information can be gathered about a newly arising disease, the earlier scientific experts and researchers would be able to procure novel treatments, and ideally, cures. This information begins with early-outbreak case reports and observational studies, where the basic characteristics of the novel disease are documented, and hence, awareness can be raised . FurtherTo the best of our knowledge, this is the first bibliometric analysis to study the worldwide COVID-19 research output. Nevertheless, there are some limitations in our study. Even though we made use of both PubMed and the WHO databases, there still might be publications that were not in our scope. We were also dependent on the indexing of the databases used, as is the case in any other bibliometric study.The outbreak of COVID-19 has caused a major threat to the international community and has raised significant public health concerns. The wide spread of this disease, along with its high rate of infectivity has incited a global demand for relevant research that can help describe the clinical and pathogenic characteristics of this illness. Observational studies related to COVID-19 can help describe the symptomatology of the disease, assess the efficiency of diagnostic tools, and establish proper management guidelines. Therapeutic trials can help discover novel treatments and come up with new curative options. Raising these concerns is essential for increasing the global research output pertaining to COVID-19.


These findings will be useful to promote educational interventions for community pharmacists regarding common infectious disease management in order to improve appropriate antibiotic use.In Thailand, antibiotics are available lawfully from community pharmacies without a prescription. Inappropriate supply of antibiotics from Thai community pharmacies to the public for common, self-limiting diseases has been reported. The study aimed to evaluate the appropriateness of antibiotics selected by community pharmacists in Thailand in response to vignettes. A cross-sectional survey of community pharmacists across Thailand was conducted using a self-administered questionnaire including nine case vignettes with three conditions, namely upper respiratory infections (URIs), acute diarrhoea and simple wounds. A total of 208 questionnaires were completed and analysed (20.8% response rate). In response to vignettes relating to URIs, 50.8% of pharmacist recommendations were not in accordance with antibiotic guidelines. Inappropriate recommendations for diarrhoea and wound cases were 20.8% and 16.7%, respectively. A higher proportion of younger pharmacists, those with less experience, Pharm. D. graduate pharmacists, employee pharmacists and those pharmacists who worked in a chain pharmacy were more likely to recommend appropriate antibiotic treatment in response to the vignettes ( In recent years, the problem of antimicrobial resistance (AMR) has increased significantly and has become a serious health care issue worldwide ,2,3,4. GCommunity pharmacists are easily accessible to the public in many countries . Most anAntibiotic supplies without a prescription from community pharmacies have been found to have contributed to the inappropriate use of antibiotics, particularly in developing countries, even though antibiotic supply without a prescription is prohibited by the law ,16,17,18In Thailand, the Antibiotics Smart Use (ASU) Program was implemented in 2007. This programme aimed to reduce unnecessary use of antibiotics for common self-limiting conditions, including URIs, acute diarrhoea and simple wounds . In 2017Thai practice guidelines for the management of URIs, acute diarrhoea and simple wounds in community pharmacies have been published ,27. AccoA total of 211 community pharmacists out of the thousand pharmacists in the sample responded to the survey. Ninety questionnaires were returned by post; two questionnaires were excluded because of the return of a blank questionnaire. One hundred twenty-one respondents in the sample completed the questionnaire. The response rate was 20.8%. Demographic data of respondents is shown in N = 130/624) and 16.7% (N = 104/622) of antibiotic recommendations for the three diarrhoea cases  and three wound vignettes , respectively, were inappropriate  of treatment recommendations for the three URI vignettes  in recommending an antibiotic for the vignette cases, some of them suggested inappropriate antibiotic regimens.N = 100/208) of community pharmacists incorrectly recommended antibiotics for this vignette. The most commonly suggested antibiotic was amoxicillin . Other suggested antibiotics were azithromycin , roxithromycin , co-amoxiclav , clarithromycin  and erythromycin .For vignette “a”, a child who was unlikely to have a Group A Streptococcal (GAS) infection, antibiotics were not recommended by the practice guidelines. However, 48.1% (N = 74/206) of respondents suggested a teenager, vignette “b”, to have antibiotics, while antibiotics might likely be of benefit. Of 74 respondents who suggested that an antibiotic was required, 19 (25.7%) selected an antibiotic treatment as recommended by the practice guidelines, that is, amoxicillin 500 mg twice daily for ten days. Other suggested antibiotics not in accordance with guidelines were roxithromycin , azithromycin , co-amoxiclav , and co-trimoxazole .In total, 35.9% (N = 206/208) of pharmacists recommended antibiotics for the patient. However, only 22.8% (N = 47/206) of community pharmacists suggested appropriate antibiotic treatment as recommended by the practice guidelines. Amoxicillin was the most common antibiotic suggestion by respondents. However, 67 pharmacists (32.5%) stated they would supply too high a dose of amoxicillin, 1500–2000 mg per day. Almost half of participants  recommended a sub-optimal duration of amoxicillin treatment, mostly five to seven days instead of the recommended 10 days. The most common inappropriate antibiotic recommended was co-amoxiclav . In the third URI vignette (vignette “c”), antibiotic treatment would be likely to be of benefit for the patient. In this case, 99.0% (N = 208) not recommending antibiotics. Antibiotics recommended inappropriately were norfloxacin  and tetracycline .Antibiotics were not required for case “d”, diarrhoea, with 97.6% of respondents (N = 48/208) recommended antibiotics for this patient, which were nifuroxazide , co-trimoxazole , norfloxacin , azithromycin , amoxicillin , furazolidone , cefixime  and cefdinir . The other four pharmacists did not specify the name of the antibiotic.The child in case “f” was likely to have had a viral infection, based on Thai guidelines, resulting in diarrhoea, for which antibiotics were not required. In total, 23.1% of pharmacists (N = 189/208) recommended antibiotic treatment for this patient. Most of them  recommended an appropriate antibiotic treatment for shigellosis, ciprofloxacin 500 mg twice daily for 3 days  or norfloxacin 400 mg twice daily for 3-5 days . In terms of inappropriate recommendations, supplying antibiotics for longer than the recommended duration of treatment was the most common reason . Four respondents would refer the patient to consult a doctor.For case “e”, the patient was likely to have shigellosis. About ninety per cent of pharmacists  recommended antibiotic treatment for case “g” where antibiotics were not required. Antibiotics suggested for this case included dicloxacillin , cloxacillin , topical fusidic acid , topical gentamicin  and topical mupirocin .Twenty-eight respondents  did not recommend antibiotic treatment for the girl, which was the appropriate response. Antibiotics suggested for this case were dicloxacillin , cloxacillin , cephalexin , amoxicillin , co-amoxiclav  and topical fusidic acid ,For case “h”, 87.4% of participants (N = 203/208) recommended antibiotics for the infected wound scenario “i”. Most pharmacists  suggested the correct antibiotic treatment as dicloxacillin 250–500 mg four times daily  or cloxacillin 500 mg four times daily .Almost all pharmacists  scored 1/3 points on the appropriate recommendation score of URI vignettes. Most pharmacists recommended appropriate antibiotic treatment for the diarrhoea and wound vignettes. About half of participants  scored 3/3 points on the appropriate recommendation score for diarrhoea vignettes. In addition, 64.6% (N = 133/206) of participants scored 3/3 points on the appropriate recommendation score for wound vignettes.According to the responses to the vignettes, appropriate recommendation scores were calculated see . The resp = 0.01) and total recommendation score with no correlation found with other conditions. The education level of pharmacists was found to correlate with the total appropriate recommendation score. Pharmacists who graduated from a Pharm D programme were found to have a higher appropriate recommendation score than those who graduated with a BPharm and postgraduate degree (p < 0.05). The role of the pharmacist and type of community pharmacy were significantly correlated with appropriate recommendation score on antibiotic supply for URI vignettes, diarrhoea vignettes and total appropriate recommendation score. Pharmacists who were an employee or worked in a chain pharmacy were seen to have higher appropriate recommendation scores for URI vignettes, diarrhoea vignettes and for the total appropriate recommendation score. An accredited pharmacy was found to be correlated with the appropriate recommendation score of antibiotic supply for URIs vignettes (p = 0.04.)Bivariate correlations were undertaken to identify the demographic data that may be correlated with the intention-to-supply score for antibiotic treatment. The results are presented in N = 1690) of pharmacies supplied antibiotics for acute adult URIs. In addition, a study in Sri Lanka reported that 43.3% (26/60 pharmacies) of pharmacy staff supplied antibiotics for acute sore throat [Nine vignettes, comprising three URI vignettes, three diarrhoea vignettes and three wound vignettes, were used to evaluate the appropriateness of the recommended antibiotics by community pharmacists. Appropriate recommendations for URI vignettes were poor as in over half of URIs instances (50.8%), inappropriate antibiotic treatments were recommended. Less than one quarter of antibiotic recommendations for the diarrhoea (20.8%) and wound (16.7%) vignettes were inappropriate. URIs are mostly self-limiting and normally no antibiotic treatment is needed, and most guidelines do not routinely recommend antibiotic treatments for many acute URIs ,27,33,34e throat . A studye throat . Most ofThe inappropriately selected antibiotic treatments for the case vignettes in this study, across URIs, diarrhoea and simple wounds, may result from a lack of up-to-date knowledge regarding patient assessment and/or antibiotic treatments. A cross-sectional survey study in Shiraz, Iran  reportedIn this study, the findings showed that age and length of experience in community pharmacy correlated with the intention to supply appropriate antibiotics. Higher proportions of younger pharmacists and pharmacists with less experience in community pharmacy stated an appropriate antibiotic. A similar association was also reported in a previous study: it was reported that greater practice experience in community pharmacy potentially increased the likelihood of inappropriate antibiotic use in Southern Thailand . LikewisThe highest education level of pharmacists was also shown to be associated with the appropriateness of antibiotic supply. Pharmacists who graduated from a PharmD programme intended to supply antibiotics more appropriately than those who graduated with a BPharm. This is possibly the result of pharmacy education programme reforms in Thailand, transitioning from a 5-year bachelor’s degree programme to a 6-year Pharm D programme, which comes with enhanced clinical practice in the sixth year of study ,42. In aThe findings also showed that pharmacy owners were more likely to state that they would supply inappropriate antibiotics, as were community pharmacists who worked in independent pharmacies. Similarly, a cross-sectional survey study conducted in Bangkok and Chonburi in Thailand in 2017 found that pharmacists who worked in a chain pharmacy have more knowledge regarding antibiotic use than others who worked in an independent pharmacy . This isThe Community Pharmacy Accreditation Project in Thailand was introduced in 2002 to ensure the delivery of high-quality pharmaceutical care by community pharmacies in Thailand. Surprisingly, accreditation status of pharmacy was found not to be significantly associated with the appropriate supply of antibiotics. Similar results were found in a mixed-methods study using observation followed by semi-structured interview in thirty community pharmacies in Vietnam. They found that there was no significant difference between Good Pharmacy Practice (GPP) certified pharmacies and non-GPP certified pharmacies regarding antibiotic supply practice . The finOur findings revealed that many community pharmacists may lack up-to-date knowledge towards antibiotic use and antimicrobial susceptibility in Thailand. Therefore, education and training of community pharmacists regarding infectious disease management is needed. The WHO suggested establishing AMR as a core component of professional education, training, certification and development for the health sectors, including community pharmacies .This is the first cross-sectional survey about antibiotic supply from community pharmacists in Thailand recruited across Thailand. There are three key limitations to the findings of this study. Firstly, the findings may not represent the wider practice and views of community pharmacists in Thailand. However, systematic random sampling was used to recruit community pharmacists for the postal survey. The study sample was diverse in terms of gender, age, length of experience, type of pharmacy and accreditation status of pharmacy and was from different regions of the country. Secondly, as this was a self-administered questionnaire-based study, there is the possibility that participants may have over-reported desirable practices or views or under-reported undesirable practices or views. As a result, the appropriateness of antibiotic supply by community pharmacists found in this study may represent a more favourable picture than might actually be the situation. Thirdly, the study asked about the intention to supply, rather than measuring actual supply.A cross-sectional survey using a self-completed questionnaire with Thai community pharmacists was used in this study. The questionnaires were distributed to selected community pharmacies by post. The data were collected between October and December 2019.Stratified random sampling was conducted to recruit community pharmacies from all regions, each with different cultural, socio-economic and socio-demographic characteristics. To classify strata, firstly, community pharmacies were grouped based on their location into the four regions, namely central Thailand, Northern Thailand, Northeastern Thailand and Southern Thailand. Then, community pharmacies in each regional area were divided into three groups based on the population size of the province in which the pharmacy was located. Quota sampling was also used in order to recruit a reasonable number of accredited pharmacies to the study. A ratio of approximately 1:2 accredited pharmacies:non-accredited pharmacies was used See .Participants’ recommended antibiotic treatments were established through the use of vignettes. Nine vignettes, comprising three URI vignettes, three diarrhoea vignettes and three wound vignettes were included. Case vignettes were created based on a review of the literature and of Thai antibiotic practice guidelines ,27. The Survey participants were asked if they would recommend antibiotics when the patient or caregiver was to present at their pharmacy with the specified symptoms and the person presenting did not ask for a specific medicine. The pharmacist participants were asked to indicate the name of the antibiotic they would recommend, dosage regimen and duration of treatment, or other option, including no supply. For each vignette, pharmacists were asked to consider that the person was able to afford the cost of medicines, and in each case, the person with symptoms had no comorbidity or undiagnosed underlying disease, used no other medication and had no history of drug allergy or intolerance. The appropriateness of antibiotic treatment indicated by community pharmacists was assessed based on the Thai practice guidelines for community pharmacists ,27.The appropriate recommendation scores were calculated according to the appropriateness of recommended antibiotic treatments as intended by community pharmacists. The scores were used as a dependent variable to identify factors influencing the appropriateness of antibiotic supply. The correct answers were identified according to the Rational Drug Use in community pharmacy, Thailand guideline . CorrectData analyses were performed using IBM SPSS statistics 25. As the study consisted of two different methods of response using the same questionnaire, there were two data sets based on the survey . The homogeneity of variance and the difference between the demographic data of two datasets were tested to see if the survey data from the two methods could be combined. Kruskal–Wallis H test was used to determine if there were statistically significant differences between the two methods of responding to the survey with regards to the variance of community pharmacists’ appropriate recommendation score of antibiotic supply. Then, a chi-square test  or a Mann–Whitney U test (for other variables) was used to test the difference between the two groups by looking at the demographic data.Nonparametric tests were used to compare the data from both surveys due to the appropriate recommendation score not being normally distributed. Bivariate analyses were conducted to identify the factors that correlated with pharmacists’ appropriate recommendation score using Pearson’s correlation (for continuous variables) or Spearman’ rank test .Ethical approval was obtained from Cardiff University School of Pharmacy and Pharmaceutical Sciences Ethics Committee (in English 1819-22) and the Research Ethics Committee of Ubon Ratchathani University, Thailand (in Thai UBU-REC-28/2562).Factors such as age, length of experience in community pharmacy, highest education level and employment status were found to be associated with the appropriateness of antibiotic supply. These findings may relate to a lack of up-to-date knowledge by community pharmacists regarding treatment and diagnosis of infectious disease.