Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Transdifferentiation”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 829 records · Page 46Linked to original sources
PubMed2003

Simulated microgravity suppresses osteoblast phenotype, Runx2 levels and AP-1 transactivation.

Conditions of disuse such as bed rest, space flight, and immobilization result in decreased mechanical loading of bone, which is associated with reduced bone mineral density and increased fracture risk. Mechanisms involved in this process are not well understood but involve the suppression of osteoblast function. To elucidate the influence of mechanical unloading on osteoblasts, a rotating wall vessel (RWV) was employed as a ground based model of simulated microgravity. Mouse MC3T3-E1 osteoblasts were grown on microcarrier beads for 14 days and then placed in the RWV for 24 h. Consistent with decreased bone formation during actual spaceflight conditions, alkaline phosphatase and osteocalcin expression were decreased by 80 and 50%, respectively. In addition, runx2 expression and AP-1 transactivation, key regulators of osteoblast differentiation and bone formation, were reduced by more than 60%. This finding suggests that simulated microgravity could promote dedifferentiation and/or transdifferentiation to alternative cell types; however, markers of adipocyte, chondrocyte, and myocyte lineages were not induced by RWV exposure. Taken together, our results indicate that simulated microgravity may suppress osteoblast differentiation through decreased runx2 and AP-1 activities.

3T3 Cells↗
PubMed2003

Differentiation potential of a mouse bone marrow stromal cell line.

In order to study osteoblast differentiation we subcloned a cell derived from a mouse a bone marrow stromal cell line, Kusa O, and obtained a number of clones representative of three different phenotypes. One that neither differentiated into osteoblasts nor into adipocytes, a second that differentiated into osteoblasts but not adipocytes, and a third that differentiated into both osteoblasts and adipocytes. Four subclones were selected for further characterization according to their ability to mineralize and/or differentiate into adipocytes. The non-mineralizing clone had no detectable alkaline phosphatase activity although some alkaline phosphatase mRNA was detected after 21 days in osteoblast differentiating medium. Alkaline phosphatase activity and mRNA in the three mineralizing clones were comparable with the parent clones. Osteocalcin mRNA and protein levels in the non-mineralizing clone were low and non-detectable, respectively, while both were elevated in the parent cells and mineralizing subclones after 21 days in differentiating medium. PTH receptor mRNA and activity increased in the four subclones and parent cells with differentiation. mRNA for two other osteoblast phenotypic markers, osteopontin and bone sialoprotein, were similarly expressed in the parent cells and subclones while mRNAs for the transcription factors, Runx2 and osterix, were detectable in both parent and subclone cells. Runx2 was unchanged with differentiation while osterix was increased. Interestingly, PPARgamma mRNA expression did not correlate with cell line potential to differentiate into adipocytes. Indian hedgehog mRNA and its receptor (patched) mRNA levels both increased with differentiation while mRNA levels of the Wnt pathway components beta-catenin and dickkopf also increased with differentiation. Although we have focussed on characterizing these clones from the osteoblast perspective it is clear that they may be useful for studying both osteoblast and adipocyte differentiation as well as their transdifferentiation.

Adipocytes↗
PubMed2004

Modulation of prostate cancer growth in bone microenvironments.

Bone remains one of the major sites, and most lethal host organs, for prostate cancer metastasis. Prostate cell spread and establishment in bone depends on multiple reciprocal modifications of bone stromal and epithelial cancer cell behaviors. This review focuses on recent advances in the characterization of cell-cell and cell-matrix interplay, effects on cell growth, adhesion and invasion, and several therapeutic possibilities for co-targeting prostate cancer cells and bone stroma. We address the topic from three main perspectives: (1) the normal and aging bone stromal environment, (2) the "reactive" bone stromal environment, and (3) the cancerous prostate epithelial cells themselves. First, normal, and especially aging, bones provide uniquely rich and "fertile soil" for roaming cancer cells. The interactions between prostate cancer cells and insoluble extracellular matrices, soluble growth factors, and/or sex steroid hormones trigger bone remodeling, through increased osteoclastogenesis and furthur matrix metalloproteinase activity. Second, after cancer cell arrival and establishment in the bone, host stromal cells respond, becoming "reactive" in a process again involving extracellular matrix remodeling, together with growth factor and steroid receptor signaling this process ultimately enhances cancer cell migration, stromal transdifferentiation, and invasion of the cancer tissues by stromal, inflammatory, and immune-responsive cells. Third, prostate cancer cells also respond to supportive bone microenvironments, where soluble and matrix-associated molecules affect cancer cell growth and gene expression, especially altering cancer cell surface receptor and integrin-mediated cell signaling. We discuss both integrin cell-matrix and gap junctional cell-cell communication between cancer cells and their microenvironments during prostate cancer progression.

Animals↗
PubMed2005

TGF-beta control of cell proliferation.

This article focuses on recent findings that the type V TGF-beta receptor (TbetaR-V), which co-expresses with other TGF-beta receptors (TbetaR-I, TbetaR-II, and TbetaR-III) in all normal cell types studied, is involved in growth inhibition by IGFBP-3 and TGF-beta and that TGF-beta activity is regulated by two distinct endocytic pathways (clathrin- and caveolar/lipid-raft-mediated). TGF-beta is a potent growth inhibitor for most cell types, including epithelial and endothelial cells. The signaling by which TGF-beta controls cell proliferation is not well understood. Many lines of evidence indicate that other signaling pathways, in addition to the prominent TbetaR-I/TbetaR-II/Smad2/3/4 signaling cascade, are required for mediating TGF-beta-induced growth inhibition. Recent studies revealed that TbetaR-V, which is identical to LRP-1, mediates IGF-independent growth inhibition by IGFBP-3 and mediates TGF-beta-induced growth inhibition in concert with TbetaR-I and TbetaR-II. In addition, IRS proteins and a Ser/Thr-specific protein phosphatase(s) are involved in the TbetaR-V-mediated growth inhibitory signaling cascade. The TbetaR-V signaling cascade appears to cross-talk with the TbetaR-I/TbetaR-II, insulin receptor (IR), IGF-I receptor (IGF-IR), integrin and c-Met signaling cascades. Attenuation or loss of the TbetaR-V signaling cascade may enable carcinoma cells to escape from TGF-beta growth control and may contribute to the aggressiveness and invasiveness of these cells via promoting epithelial-to-mesenchymal transdifferentiation (EMT). Finally, the ratio of TGF-beta binding to TbetaR-II and TbetaR-I is a signal controlling TGF-beta partitioning between two distinct endocytosis pathways and resultant TGF-beta responsiveness. These recent studies have provided new insights into the molecular mechanisms underlying TGF-beta-induced cellular growth inhibition, cross-talk between the TbetaR-V and other signaling cascades, the signal that controls TGF-beta responsiveness and the role of TbetaR-V in tumorigenesis.

Animals↗
PubMed2006

Are there morally acceptable alternatives to blastocyst derived ESC?

ESC derivation, use and SCNT have raised many moral and ethical issues. In this opinion piece I have focused on the argument that morally less ambiguous alternatives to ESC derived from the ICM of blastocysts exist. These possibilities range from using multiple adult stem cell populations each of which is uniquely suited for a particular disease target or identifying adult ESC-like populations, using transdifferentiated ESC-like cells or alternate methods of deriving ESC. I suggest that while it is important to support such efforts, current results do not provide sufficient compelling data to allow one to stop the use of ESC and perhaps adult cells will never be a reliable alternative. All options need to be fully explored and decisions need to be made with scientific rigor and respect for each individual's moral compass.

Aging↗
PubMed1989

Stochastic model for mast cell proliferation in culture of murine peritoneal cells.

We recently identified two types of mast cell colonies derived from murine peritoneal cells: type 1 and type 2. Type 1 mast cell colonies consisted of berberine sulfate(+)- safranin(+) connective tissue-type mast cells (CTMC) and were derived from mature CTMC in the heaviest fraction obtained by Percoll density gradient centrifugation. In contrast, type 2 mast cell colonies consisted of alcian blue(+)- berberine sulfate(-)- safranin(-) mucosal mast cells (MMC) and were derived from immature progenitors in low density fractions. We replated a total of 60 type 1 and 60 type 2 mast cell colonies and examined their capability for producing secondary colonies. Although all of the primary colonies yielded secondary colonies, the replating efficiencies of individual colonies varied over a wide range. Cumulative distributions of secondary colonies from both type 1 and type 2 primary colonies could be fitted well by gamma distributions obtained by computer simulation. These findings are in agreement with the stochastic model for CTMC- and MMC proliferation. Cytological analyses of secondary colonies from primary type 1 colonies revealed heterogeneous distributions of alcian blue(+)- safranin(-)- berberine sulfate(-) mast cells, suggesting that transdifferentiation from mature CTMC to safranin(-)- berberine sulfate(-) mast cells is also governed by stochastic mechanisms.

Animals↗
PubMed2005

3,5,3'-Triiodo-L-thyronine enhances the differentiation of a human pancreatic duct cell line (hPANC-1) towards a beta-cell-Like phenotype.

The thyroid hormone, 3,5,3'-Triiodo-L-thyronine (T3), is essential for growth, differentiation, and regulation of metabolic functions in multicellular organisms, although the specific mechanisms of this control are still unknown. In this study, treatment of a human pancreatic duct cell line (hPANC-1) with T3 blocks cell growth by an increase of cells in G(0)/G(1) cell cycle phase and enhances morphological and functional changes as indicated by the marked increase in the synthesis of insulin and the parallel decrease of the ductal differentiation marker cytokeratin19. Expression analysis of some of the genes regulating pancreatic beta-cell differentiation revealed a time-dependent increase in insulin and glut2 mRNA levels in response to T3. As last step of the acquisition of a beta-cell-like phenotype, we present evidence that thyroid hormones are able to increase the release of insulin into the culture medium. In conclusion, our results suggest, for the first time, that thyroid hormones induce cell cycle perturbations and play an important role in the process of transdifferentiation of a human pancreatic duct line (hPANC-1) into pancreatic-beta-cell-like cells. These findings have important implications in cell-therapy based treatment of diabetes and may provide important insights in the designing of novel therapeutic agents to restore normal glycemia in subjects with diabetes.

Carcinoma, Pancreatic Ductal↗
PubMed2006

SOCS-2 interferes with myotube formation and potentiates osteoblast differentiation through upregulation of JunB in C2C12 cells.

Suppressor of cytokine signaling (SOCS)-2 regulates normal postnatal growth and its deficiency in mice causes gigantism with increased bone length and proportional enlargement in skeletal muscles. Using C2C12 mesenchymal precursor cell line as a model, we investigated a possible role of SOCS-2 in the differentiation process of mesenchymal precursors. Stable transfection of SOCS-2 into C2C12 cells resulted in the acceleration of proliferation and survival, and inhibition of spontaneous myotube formation. In addition, SOCS-2 potentiated bone morphogenic protein (BMP)-induced transdifferentiation of C2C12 cells into osteoblast phenotypes. These effects of SOCS-2 on C2C12 cells differed strikingly from that of SOCS-1, another member of SOCS family, and its mechanisms were evaluated. SOCS-2 did not alter BMP-induced phosphorylation and nuclear accumulation of Smad1, nor the expression of inhibitory-Smads mRNA. However, SOCS-2 enhanced BMP-induced transcriptional activation of the Smad-responsive reporter gene, suggesting that the action of SOCS-2 is exerted at the transcriptional level. Interestingly, SOCS-2 overexpression in C2C12 cells increased the endogenous JunB protein, one of the key transcriptional factors in the control of BMP/Smad signaling responsiveness. In addition, the proteasome inhibitor enhanced JunB protein expression in C2C12 cells. Moreover, we found that SOCS-2 reduced JunB ubiquitination in COS-7 cells. Although SOCS-2 is a modulator of growth hormone (GH) signaling, the upregulation of JunB by SOCS-2 did not require GH signaling. Taken together, these results suggest that SOCS-2 positively regulates endogenous JunB protein expression in C2C12 cells through inhibition of JunB destabilization by the ubiquitin-proteasome pathway, and thereby regulates the cell fate of mesenchymal precursors.

Alkaline Phosphatase↗
PubMed2002

Role of cell cycle molecules in the pathophysiology of glomerular epithelial cells.

The postmitotic characteristics of podocytes are the basis for the structural assembly and central function (filtration) of the glomeruli. Persistent cell cycle quiescence is required for stability in these cells. In cells of the podocyte lineage, transdifferentiation from the metanephric mesenchyme to mature podocytes is closely associated with tight regulation of the expression of cell cycle molecules. This cell cycle control in podocytes acts as a safeguard for the glomeruli; however, deregulation of this system might result in structural deterioration. This article focuses on the expression of cell cycle molecules in podocyte differentiation and pathology.

Cell Cycle Proteins↗
PubMed2001

Sex reversal and aromatase in the European pond turtle: treatment with letrozole after the thermosensitive period for sex determination.

In the European pond turtle (Emys orbicularis), gonadal sex differentiation is temperature-dependent. The temperature sensitive period (TSP) of gonadogenesis lies between stages 16 and 22 of embryonic development. Previous studies have shown that embryos incubated at 30 degrees C, a temperature yielding 100% phenotypic females, can be sex reversed by treatments with an aromatase inhibitor administered during TSP or even somewhat after TSP (as of stage 22+). The goal of the present study was to determine whether the ovary still retains male potential at later stages of embryonic development and whether the induced male characters persist after hatching. For this purpose, eggs of E. orbicularis were treated with letrozole, a nonsteroidal aromatase inhibitor, at or as of stages 23, 24 or 25, then gonadal aromatase activity in each individual and the related gonadal structure were studied at hatching (stage 26) and for one year after hatching. Two kinds of treatments were carried out: 1) repeated applications of 10 microg of letrozole in ethanolic solution onto the eggshell; and 2) a single injection of 10 microg of letrozole in olive oil. Similar results were obtained with either application or injection of the aromatase inhibitor. In treatments as of or at stage 23, individuals with gonadal aromatase activity lower than 20 fmoles/hour/gonad had ovotestes, i.e., 22% of the treated individuals. At hatching, the inner part of these ovotestes contained testicular cords and also mixed lacunae presenting various degrees of transdifferentiation of the epithelium into a Sertolian epithelium. The cortex was maintained, although some germ cells degenerated within it. These processes continued after hatching. However, at 12 months, gonads were still ovotestes displaying some follicles with a growing oocyte in the remaining parts of the cortex. In treatments as of or at stages 24 or 25, only a few individuals were masculinized. One had ovotestes; in others, the cortex was absent in some parts and when it was present oocytes were degenerating. These results show that in the European pond turtle, differentiation of ovotestes from ovaries can be induced by treatment with an aromatase inhibitor starting at late stages of embryonic development (between the end of TSP and hatching), although such differentiation is less frequent as embryonic development proceeds. Sex reversal persists for at least one year after hatching. J. Exp. Zool. 290:490-497, 2001.

Animals↗
PubMed1978

Insect cellular differentiation. I. Redifferentiation of the adult labial glands of Manduca sexta (Lepidoptera).

With the exception of a few primitive ephemeropterans, winged insects do not normally molt as adults, although they can molt under appropriate hormonal conditions. We investigated this supernumerary molt process by taking adult labial glands (from Manduca sexta), cutting them into fragments of a single cell type, implanting them into individual freshly molted pupae, and then recovering them from pharate adult and adult hosts. The appearance and function of these fragments were compared to those of normally metamorphosing glands. Based on these criteria: (1) the supernumerary adult molt involved cellular de- and re-differentiation and (2) no transdifferentiation occurred, between the separate adult gland cell types.

Animals↗
PubMed2003

Tissue interactions and lens-forming competence in the outer cornea of larval Xenopus laevis.

After lentectomy through the pupillary hole, the outer cornea of larval Xenopus laevis can undergo transdifferentiation to regenerate a new lens. This process is elicited by inductive factor(s) produced by the neural retina and accumulated into the vitreous chamber. During embryogenesis, the outer cornea develops from the outer layer of the presumptive lens ectoderm (PLE) under the influence of the eye cup and the lens. In this study, we investigated whether the capacity of the outer cornea to regenerate a lens is the result of early inductive signals causing lens-forming bias and lens specification of the PLE, or late inductive signals causing cornea formation or both signals. Fragments of larval epidermis or cornea developed from ectoderm that had undergone only one kind of inductive signals, or both kinds of signals, or none of them, were implanted into the vitreous chamber of host larvae. The regeneration potential and the lens-forming transformations of the implants were tested using an antisense probe for pax6 as an earlier marker of lens formation and a monoclonal antibody anti-lens as a definitive indicator of lens cell differentiation. Results demonstrated that the capacity of the larval outer cornea to regenerate a lens is the result of both early and late inductive signals and that either early inductive signals alone or late inductive signals alone can elicit this capacity.

Animals↗
PubMed2006

Experimental analysis of lens-forming capacity in Xenopus borealis larvae.

Previously, the only anuran amphibians known to have the capacity to regenerate a lens after lentectomy were Xenopus laevis and Xenopus tropicalis. This regeneration process occurs during the larval life through transdifferentiation of the outer cornea promoted by inductive factors produced by the retina and accumulated inside the vitreous chamber. However, the capacity of X. tropicalis to regenerate a lens is much lower than that of X. laevis. This study demonstrates that Xenopus borealis, a species more closely related to X. laevis than to X. tropicalis, is not able to regenerate a lens after lentectomy. Nevertheless, some morphological modifications corresponding to the first stages of lens regeneration in X. laevis were observed in the outer cornea of X. borealis. This suggested that in X borealis the regeneration process was blocked at early stages. Results from histological analysis of X. borealis and X. laevis lentectomized eyes and from implantation of outer cornea fragments into the vitreous and anterior chambers demonstrated that: (i) in X. borealis eye, the lens-forming competence in the outer cornea and inductive factors in the vitreous chamber are both present, (ii) no inhibiting factors are present in the anterior chamber, the environment where lens regeneration begins, (iii) the inability of X. borealis to regenerate a lens after lentectomy is due to an inhibiting action exerted by the inner cornea on the spreading of the retinal factor from the vitreous chamber towards the outer cornea. This mechanical inhibition is assured by two distinctive features of X. borealis eye in comparison with X. laevis eye: (i) a weaker and slower response to the retinal inducer by the outer cornea; (ii) a stronger and faster healing of the inner cornea. Unlike X. tropicalis and similar to X. laevis, in X. borealis the competence to respond to the retinal factor is not restricted to the corneal epithelium but also extends to the pericorneal epidermis.

Animals↗
PubMed2003

Preservation of hematopoietic properties in transplanted bone marrow cells in the brain.

Recent studies have described the possible transdifferentiation of bone marrow cells (BMC) into neurons and glia when they migrate to the brain. However, we have reported that some immature BMC migrating into the brain parenchyma after bone marrow transplantation express early hematopoietic markers but not neural or glial markers. The present study further characterizes transplanted BMC that migrate to the brain. Double immunolabeling confirmed that BMC migrating to the brain expressed hematopoietic but not neural markers, such as nestin, microtubule-associated protein-2 and glial fibrillary acidic protein, even 4 and 18 weeks after bone marrow transplantation. BMC that expressed green fluorescent protein also expressed hematopoietic but not neural markers when cultured with mixed brain cells according to double immunolabeling and single-cell dissection using a laser. Analysis of the DNA content indicated that most of the migrated BMC were arrested at the G0/G1 phase, and aneuploidy or tetraploidy was undetectable. Thus, BMC that migrate to the brain probably have preserved hematopoietic properties under physiological conditions.

Animals↗
PubMed2004

Reevaluation of in vitro differentiation protocols for bone marrow stromal cells: disruption of actin cytoskeleton induces rapid morphological changes and mimics neuronal phenotype.

Bone marrow stromal cells (MSC), which represent a population of multipotential mesenchymal stem cells, have been reported to undergo rapid and robust transformation into neuron-like phenotypes in vitro following treatment with chemical induction medium including dimethyl sulfoxide (DMSO; Woodbury et al. [2002] J. Neurosci. Res. 96:908). In this study, we confirmed the ability of cultured rat MSC to undergo in vitro osteogenesis, chondrogenesis, and adipogenesis, demonstrating differentiation of these cells to three mesenchymal cell fates. We then evaluated the potential for in vitro neuronal differentiation of these MSC, finding that changes in morphology upon addition of the chemical induction medium were caused by rapid disruption of the actin cytoskeleton. Retraction of the cytoplasm left behind long processes, which, although strikingly resembling neurites, showed essentially no motility and no further elaboration during time-lapse studies. Similar neurite-like processes were induced by treating MSC with DMSO only or with actin filament-depolymerizing agents. Although process formation was accompanied by rapid expression of some neuronal and glial markers, the absence of other essential neuronal proteins pointed toward aberrantly induced gene expression rather than toward a sequence of gene expression as is required for neurogenesis. Moreover, rat dermal fibroblasts responded to neuronal induction by forming similar processes and expressing similar markers. These studies do not rule out the possibility that MSC can differentiate into neurons; however, we do want to caution that in vitro differentiation protocols may have unexpected, misleading effects. A dissection of molecular signaling and commitment events may be necessary to verify the ability of MSC transdifferentiation to neuronal lineages.

Actin Cytoskeleton↗
PubMed2004

Regeneration of the inner ear as a model of neural plasticity.

The publication of a paper entitled "Direct transdifferentiation gives rise to the earliest new hair cells in regenerating avian auditory epithelium" in the Journal of Neuroscience Research offers the opportunity to call attention to a well-developed line of research on the auditory receptor of birds, which should be of interest to students of regeneration and plasticity of the mature nervous system in higher vertebrates, including mammals. Although hair cell proliferation normally stops before hatching, destruction of the auditory receptors of the chicken may be followed by complete regeneration of hair cells. Most of the new hair cells arise from a new wave of proliferation, but Roberson et al. show that about one-third of the new hair cells are formed without undergoing cell division and thus may differentiate from so-called supporting cells or cells with an "intermediate morphology." This finding suggests some models for regeneration of this neuroepithelium, including the possibility that mature supporting cells could transform directly into hair cells. The present Mini-Review discusses some of the models for neural regeneration that future studies might address in the light of our current knowledge and the new report. The possibility is raised that transitional forms of hair cell and supporting cell precursors may reside in the inner ear in a quiescent state until stimulated by damage.

Animals↗
PubMed2007

2', 3'-Cyclic nucleotide 3'-phosphodiesterase cells derived from transplanted marrow stromal cells and host tissue contribute to perineurial compartment formation in injured rat spinal cord.

Transdifferentiation of transplanted marrow stromal cells (MSCs) and reactive changes of glial cells in a completely transected rat spinal cord were examined. Marrow stromal cells exhibited 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNP) at the plasma membrane and this has allowed their identification after transplantation by immunoelectron microscopy. In the control rats, the lesion site showed activated microglia/neural macrophages and some elongated cells, whose cytoplasm was immunoreactive for CNP. Cells designated as CNP1 and apparently host-derived expressed CXCR4. In experimental rats receiving MSCs transplantation, CNP1 cells were increased noticeably. This was coupled with the occurrence of a different subset of CNP cells whose plasma membrane was CNP-immunoreactive and expressed CXCR4. These cells, designated as CNP2, enclosed both myelinated and unmyelinated neurites thus assuming a spatial configuration resembling that of Schwann cells. A remarkable feature was the extensive ramifications of CNP1 cells with long filopodia processes delineating the CNP2 cells and their associated neurites, forming many perineurial-like compartments. Present results have shown that CNP2 cells considered to be MSCs-derived can transform into cells resembling Schwann cells based on their spatial relation with the regenerating nerve fibers, whereas the CNP1 glial cells participate in formation of perineurial compartments, probably serving as conduits to guide the nerve fiber growth. The chemotactic migration of CNP cells either derived from host tissue or MSCs bearing CXCR4 may be attracted by stromal derived factor-1alpha (SDF-1alpha) produced locally. The coordinated cellular interaction between transplanted MSCs and local glial cells may promote the growth of nerve fibers through the lesion site.

2',3'-Cyclic-Nucleotide Phosphodiesterases↗
PubMed1998

Programmed removal of chondrocytes during endochondral fracture healing.

This investigation tested the hypothesis that the removal of chondrocytes during endochondral fracture healing involves an ordered process of programmed cell death. To accomplish this, unilateral closed fractures were created in the femora of 36 Sprague-Dawley rats. The rats were killed in groups of four on days 1, 3, 7, 14, 21, 28, 42, 49, and 56 after fracture. The femora were embedded in paraffin and tested for expression of specific markers of fragmented DNA with use of a terminal deoxyuridyl transferase-mediated deoxyuridine triphosphate-biotin nick end labeling (TUNEL) technique. To determine the potential for transdifferentiation of chondrocytes to osteoblasts, calluses were also hybridized to detect expression of osteocalcin mRNA. Cell proliferation was assessed by an immunohistochemical detection method for proliferating cell nuclear antigen. A separate group of four rats was killed on day 28 to represent the later stage of the endochondral ossification, and the calluses were examined for cellular morphology with transmission electron microscopy. The results showed a coordination in both time and space of the activities of cellular proliferation and programmed cell death. Cell proliferation was most active in the earlier phases of fracture healing (days 1 through 14), although TUNEL expression was apparent in hypertrophic chondrocytes on day 14 after fracture and persisted until day 28. In the later stages of fracture healing (days 14 through 28), proliferating cell nuclear antigen was no longer synthesized in hard callus (intramembranous bone) and cell removal was the dominant activity in soft callus chondrocytes. Expression of osteocalcin mRNA was detected in osteoblasts but not in hypertrophic chondrocytes or in any other nonosteoblastic cell type. These findings support the hypothesis that the removal of chondrocytes during endochondral fracture healing is part of an ordered transition of tissue types in which the cellular mechanisms are genetically programmed to involve proliferation, maturation, and apoptotic cell death.

Animals↗