RESUMEN
Mercer University School of Medicine utilizes a problem-based learning (PBL) curriculum for educating medical students in the basic clinical sciences. In 2014, an adjustment was piloted that enabled PBL cases to align with their corresponding cadaver dissection that reviewed the content of anatomy contained in the PBL cases. Faculty had the option of giving PBL cases in sequence with the cadaveric dissection schedule (sequential group) or maintaining PBL cases out of sequence with dissections (traditional group). During this adjustment, students' academic performances were compared. Students' perception of their own preparedness for cadaveric dissection, their perceived utility of the cadaver dissections, and free-response comments were solicited via an online survey. There were no statistically significant differences when comparing student mean examination score values between the sequential and traditional groups on both multidisciplinary examinations (79.39 ± 7.63 vs. 79.88 ± 7.31, P = 0.738) and gross anatomy questions alone (78.15 ± 10.31 vs. 79.98 ± 9.31, P = 0.314). A statistically significant difference was found between the sequential group's and traditional group's (63% vs. 29%; P = 0.005) self-perceived preparedness for cadaveric dissections in the 2017 class. Analysis of free-response comments found that students in the traditional group believed their performance in PBL group, participation in PBL group and examination performance was adversely affected when compared to students with the sequential schedule. This study provides evidence that cadaveric dissections scheduled in sequence with PBL cases can lead to increased student self-confidence with learning anatomy but may not lead to improved examination scores.
Asunto(s)
Anatomía/educación , Disección , Aprendizaje Basado en Problemas/métodos , Autoimagen , Estudiantes de Medicina/psicología , Rendimiento Académico , Adulto , Cadáver , Curriculum , Educación de Pregrado en Medicina/métodos , Femenino , Humanos , Masculino , Sistema Musculoesquelético/anatomía & histología , Estudiantes de Medicina/estadística & datos numéricos , Encuestas y Cuestionarios/estadística & datos numéricos , Adulto JovenRESUMEN
This study was designed to determine if trauma causes the release of adult-derived blastomere-like stem cells (BLSCs) from skeletal muscle into the circulating blood of adult pigs. Experimental procedures followed the guidelines of Fort Valley State University's Institutional Animal Care and Utilization Committee. Pigs were traumatized by splenectomy followed by pancreatectomy. Blood samples and skeletal muscle biopsies were taken before and after trauma. Adult-derived BLSCs were isolated from skeletal muscle and blood samples following established procedures. Nontraumatized skeletal muscle contained approximately 277 million BLSCs per gram of muscle. After trauma, skeletal muscle contained approximately 2 million BLSCs per gram of muscle. Blood taken before trauma contained approximately 22 million BLSCs per milliliter, whereas approximately 512 million BLSCs per milliliter were present within the blood after trauma. Blood values were statistically significant with a P < 0.05. This report is the first demonstration that trauma causes the release of adult-derived BLSCs from skeletal muscle into blood. Further studies are required to elucidate the roles that adult-derived BLSCs play in the response to injury and in the healing process. Surgeons must take a role in this evolving field.
Asunto(s)
Músculo Esquelético/patología , Pancreatectomía/efectos adversos , Esplenectomía/efectos adversos , Células Madre Totipotentes/patología , Heridas y Lesiones/sangre , Animales , Recuento de Células , Modelos Animales de Enfermedad , Músculo Esquelético/metabolismo , Porcinos , Heridas y Lesiones/patologíaRESUMEN
This report reviews three categories of precursor cells present within adults. The first category of precursor cell, the epiblast-like stem cell, has the potential of forming cells from all three embryonic germ layer lineages, e.g., ectoderm, mesoderm, and endoderm. The second category of precursor cell, the germ layer lineage stem cell, consists of three separate cells. Each of the three cells is committed to form cells limited to a specific embryonic germ layer lineage. Thus the second category consists of germ layer lineage ectodermal stem cells, germ layer lineage mesodermal stem cells, and germ layer lineage endodermal stem cells. The third category of precursor cells, progenitor cells, contains a multitude of cells. These cells are committed to form specific cell and tissue types and are the immediate precursors to the differentiated cells and tissues of the adult. The three categories of precursor cells can be readily isolated from adult tissues. They can be distinguished from each other based on their size, growth in cell culture, expressed genes, cell surface markers, and potential for differentiation. This report also discusses new findings. These findings include the karyotypic analysis of germ layer lineage stem cells; the appearance of dopaminergic neurons after implantation of naive adult pluripotent stem cells into a 6-hydroxydopamine-lesioned Parkinson's model; and the use of adult stem cells as transport mechanisms for exogenous genetic material. We conclude by discussing the potential roles of adult-derived precursor cells as building blocks for tissue repair and as delivery vehicles for molecular medicine.
Asunto(s)
Trasplante de Células Madre , Células Madre/citología , Células Madre/fisiología , Heridas y Lesiones/terapia , Adulto , Humanos , Cariotipificación , Mesodermo/citología , Mesodermo/fisiologíaRESUMEN
Stem cells isolated from adult tissues may be useful for autologous cell therapy in the nervous system. In the present study we tested the ability of multipotent stem cells isolated from adult muscle to survive and respond to migratory and differentiating cues when transplanted into the adult subventricular zone (SVZ). Prior to transplantation the cells were grown as spheres that expressed doublecortin, nestin, and betaIII-tubulin, as well as the mRNAs for the receptor EphA4 and the ligands ephrin B1, ephrin B2, but not ephrin B3. Four weeks after transplantation into the anterior part of the SVZ in adult rats, surviving cells were observed along the ventricular wall, in the SVZ, and in the posterior rostral migratory stream (RMS). None of these cells stained for betaIII-tubulin or doublecortin, which are molecules expressed by migrating neuroblasts, and none were present in the more rostral regions of the RMS or the olfactory bulb. However, most surviving transplanted cells were integrated into the wall of the lateral ventricle and expressed vimentin, a marker also expressed by ependymocytes. No tumors were observed 4 weeks posttransplantation. Our results suggest that multipotent stem cells isolated from adult muscle, which can be easily and safely isolated from patients and rapidly expanded ex vivo, may provide autologous vectors for the local delivery of secreted factors to the ventricles or nearby regions.
Asunto(s)
Diferenciación Celular/fisiología , Células Madre Multipotentes/citología , Músculo Esquelético/citología , Prosencéfalo/citología , Trasplante de Células Madre/métodos , Animales , Linaje de la Célula/fisiología , Movimiento Celular/fisiología , Supervivencia Celular/fisiología , Células Cultivadas , Proteína Doblecortina , Ventrículos Laterales , Masculino , Células Madre Multipotentes/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Prosencéfalo/cirugía , Ratas , Ratas Sprague-Dawley , Vimentina/metabolismoRESUMEN
Ligands of the glial cell line-derived neurotrophic factors (GDNF)-family are trophic factors for the development and survival of multiple cell types, however their effects on non-neuronal stem cells are unknown. We examined the action of neurturin on a candidate stem cell population isolated from adult skeletal muscles. When grown as spheres, these cells expressed mRNAs for GDNF, persephin, GFR-alpha2, GFR-alpha4 (neurturin receptor), and Ret. Exposure of these cells to neurturin significantly augmented cell numbers via increased cell proliferation. After addition of retinoic acid, the cells exited the cell cycle, developed thin processes, and became immunoreactive for betaIII-tubulin, while Ret mRNA expression decreased, without changes in the level of GFR-alpha2 mRNA. Neurturin induced an outgrowth of processes on these betaIII-tubulin positive cells. Neurturin may therefore be beneficial in the use of these multipotent cells isolated from adult muscles for autologous transplants in neurological applications.
Asunto(s)
Células Madre Multipotentes/citología , Células Madre Multipotentes/metabolismo , Músculo Esquelético/citología , Músculo Esquelético/metabolismo , Factores de Crecimiento Nervioso/metabolismo , Factores de Crecimiento Nervioso/farmacología , Neuronas/citología , Neuronas/metabolismo , Animales , Diferenciación Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Relación Dosis-Respuesta a Droga , Factor Neurotrófico Derivado de la Línea Celular Glial , Células Madre Multipotentes/efectos de los fármacos , Músculo Esquelético/efectos de los fármacos , Proteínas del Tejido Nervioso/metabolismo , Neuronas/efectos de los fármacos , Neurturina , Ratas , Ratas Sprague-Dawley , Tretinoina/farmacologíaRESUMEN
Adult stem cells derived from bone marrow, connective tissue, and solid organs can exhibit a range of differentiation potentials. Some controversy exists regarding the classification of mesenchymal stem cells as bona fide stem cells, which is in part derived from the limited ability to propagate true clonal populations of precursor cells. We isolated putative mesenchymal stem cells from the connective tissue of an adult rat (rMSC), and generated clonal populations via three rounds of dilutional cloning. The replicative potential of the clonal rMSC line far exceeded Hayflick's limit of 50-70 population doublings. The high capacity for self-renewal in vitro correlated with telomerase activity, as demonstrated by telomerase repeat amplification protocol (TRAP) assay. Exposure to nonspecific differentiation culture medium revealed multilineage differentiation potential of rMSC clones. Immunostaining confirmed the appearance of mesodermal phenotypes, including adipocytes possessing lipid-rich vacuoles, chondrocytes depositing pericellular type II collagen, and skeletal myoblasts expressing MyoD1. Importantly, the spectrum of differentiation capability was sustained through repeated passaging. Furthermore, serum-free conditions that led to high-efficiency smooth muscle differentiation were identified. rMSCs plated on collagen IV-coated surfaces and exposed to transforming growth factor-beta1 (TGF-beta1) differentiated into a homogeneous population expressing alpha-actin and calponin. Hence, clonogenic analysis confirmed the presence of a putative MSC population derived from the connective tissue of rat skeletal muscle. The ability to differentiate into a smooth muscle cell (SMC) phenotype, combined with a high proliferative capacity, make such a connective tissue-derived MSC population ideal for applications in vascular tissue construction.
Asunto(s)
Diferenciación Celular , Células Madre Mesenquimatosas , Proteína MioD/biosíntesis , Mioblastos/metabolismo , Telómero/metabolismo , Ingeniería de Tejidos , Adipocitos/citología , Adipocitos/metabolismo , Animales , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/genética , Diferenciación Celular/fisiología , División Celular/fisiología , Condrocitos/citología , Condrocitos/metabolismo , Colágeno Tipo II/biosíntesis , Células del Tejido Conectivo/citología , Células Madre Mesenquimatosas/citología , Células Madre Mesenquimatosas/fisiología , Mesodermo/citología , Mesodermo/metabolismo , Mioblastos/citología , Miocitos del Músculo Liso/citología , Miocitos del Músculo Liso/metabolismo , Ratas , Ratas Sprague-Dawley , Telómero/genética , Factor de Crecimiento Transformador beta/farmacologíaRESUMEN
Autologous cell therapies in neurodegenerative diseases and stroke will require an efficient generation of neuroprogenitors or neurons. We have previously shown that presumptive neural progenitors can be obtained from a candidate stem cell population isolated from adult skeletal muscle. Here we describe experimental conditions to isolate and characterize the cells with neurogenic potential from this population. Candidate stem cell population was isolated from adult skeletal muscle and expanded for selection during at least 30 cell divisions. FACS analysis revealed that this population was homogeneous with respect to CD45 (-), CD34 (-), and heterogeneous for CD90 (Thy-1) expression. The population was separated by cell sorting into three sub-populations based on CD90 expression (CD90-, CD90+, and CD90++) and each population expanded rapidly as free-floating spheres. When dissociated and plated in a neuronal differentiation medium, a large number of CD90+ cells acquired morphological characteristics of neuroprogenitors and neurons, and expressed markers of neurons but no markers of glial or muscle cells. In contrast, CD90- and CD90++ cells lacked this ability. Comparison of CD90+ and CD90- populations may be useful for studying the molecular characteristics defining the neuronal potential of stem cells from adult muscle. The selection of CD90+ expressing cells, combined with the growth conditions presented here, allows for rapid generation of a large number of cells which may be useful for autologous cell replacement therapies in the central nervous system.
Asunto(s)
Músculo Esquelético/citología , Células Madre/citología , Animales , Antígenos CD/inmunología , Diferenciación Celular , Separación Celular , Citometría de Flujo , Músculo Esquelético/inmunología , Ratas , Ratas Sprague-Dawley , Células Madre/inmunologíaRESUMEN
Tissue restoration is the process whereby multiple damaged cell types are replaced to restore the histoarchitecture and function to the tissue. Several theories have been proposed to explain the phenomenon of tissue restoration in amphibians and in animals belonging to higher orders. These theories include dedifferentiation of damaged tissues, transdifferentiation of lineage-committed progenitor cells, and activation of reserve precursor cells. Studies by Young et al. and others demonstrated that connective tissue compartments throughout postnatal individuals contain reserve precursor cells. Subsequent repetitive single cell-cloning and cell-sorting studies revealed that these reserve precursor cells consisted of multiple populations of cells, including tissue-specific progenitor cells, germ-layer lineage stem cells, and pluripotent stem cells. Tissue-specific progenitor cells display various capacities for differentiation, ranging from unipotency (forming a single cell type) to multipotency (forming multiple cell types). However, all progenitor cells demonstrate a finite life span of 50 to 70 population doublings before programmed cell senescence and cell death occurs. Germ-layer lineage stem cells can form a wider range of cell types than a progenitor cell. An individual germ-layer lineage stem cell can form all cells types within its respective germ-layer lineage (i.e., ectoderm, mesoderm, or endoderm). Pluripotent stem cells can form a wider range of cell types than a single germ-layer lineage stem cell. A single pluripotent stem cell can form cells belonging to all three germ layer lineages. Both germ-layer lineage stem cells and pluripotent stem cells exhibit extended capabilities for self-renewal, far surpassing the limited life span of progenitor cells (50-70 population doublings). The authors propose that the activation of quiescent tissue-specific progenitor cells, germ-layer lineage stem cells, and/or pluripotent stem cells may be a potential explanation, along with dedifferentiation and transdifferentiation, for the process of tissue restoration. Several model systems are currently being investigated to determine the possibilities of using these adult quiescent reserve precursor cells for tissue engineering.
Asunto(s)
Diferenciación Celular/fisiología , Estratos Germinativos/citología , Células Madre Pluripotentes/citología , Regeneración/fisiología , Ingeniería de Tejidos , Factores de Transcripción , Animales , Antígenos de Superficie/metabolismo , Huesos/metabolismo , Bovinos , Linaje de la Célula/fisiología , Senescencia Celular/fisiología , Tejido Conectivo/fisiología , Proteínas de Unión al ADN/metabolismo , Diabetes Mellitus/terapia , Embrión de Mamíferos , Embrión no Mamífero , Extremidades/fisiología , Terapia Genética , Trasplante de Células Madre Hematopoyéticas , Humanos , Músculo Esquelético/metabolismo , Músculo Esquelético/trasplante , Infarto del Miocardio/terapia , Miogenina/metabolismo , Enfermedades Neurodegenerativas/terapia , Factor 3 de Transcripción de Unión a Octámeros , Células Madre Pluripotentes/trasplante , Ratas , Telomerasa/metabolismo , Urodelos/crecimiento & desarrollo , Urodelos/fisiologíaRESUMEN
Undifferentiated cells have been identified in the prenatal blastocyst, inner cell mass, and gonadal ridges of rodents and primates, including humans. After isolation these cells express molecular and immunological markers for embryonic cells, capabilities for extended self-renewal, and telomerase activity. When allowed to differentiate, embryonic stem cells express phenotypic markers for tissues of ectodermal, mesodermal, and endodermal origin. When implanted in vivo, undifferentiated noninduced embryonic stem cells formed teratomas. In this report we describe a cell clone isolated from postnatal rat skeletal muscle and derived by repetitive single-cell clonogenic analysis. In the undifferentiated state it consists of very small cells having a high ratio of nucleus to cytoplasm. The clone expresses molecular and immunological markers for embryonic stem cells. It exhibits telomerase activity, which is consistent with its extended capability for self-renewal. When induced to differentiate, it expressed phenotypic markers for tissues of ectodermal, mesodermal, and endodermal origin. The clone was designated as a postnatal pluripotent epiblastic-like stem cell (PPELSC). The undifferentiated clone was transfected with a genomic marker and assayed for alterations in stem cell characteristics. No alterations were noted. The labeled clone, when implanted into heart after injury, incorporated into myocardial tissues undergoing repair. The labeled clone was subjected to directed lineage induction in vitro, resulting in the formation of islet-like structures (ILSs) that secreted insulin in response to a glucose challenge. This study suggests that embryonic-like stem cells are retained within postnatal mammals and have the potential for use in gene therapy and tissue engineering.
Asunto(s)
Ensayo de Unidades Formadoras de Colonias/métodos , Células Madre Pluripotentes/citología , Células Madre Pluripotentes/fisiología , Animales , Animales Recién Nacidos , Masculino , Ratas , Ratas Endogámicas WF , Ratas Sprague-Dawley , Células Madre/citología , Células Madre/fisiologíaRESUMEN
Development of a multicellular organism is accomplished through a series of events that are preprogrammed in the genome. These events encompass cellular proliferation, lineage commitment, lineage progression, lineage expression, cellular inhibition, and regulated apoptosis. The sequential progression of cells through these events results in the formation of the differentiated cells, tissues, and organs that constitute an individual. Although most cells progress through this sequence during development, a few cells leave the developmental continuum to become reserve precursor cells. The reserve precursor cells are involved in the continual maintenance and repair of the tissues and organs throughout the life span of the individual. Until recently it was generally assumed that the precursor cells in postnatal individuals were limited to lineage-committed progenitor cells specific for various tissues. However, studies by Young, his colleagues, and others have demonstrated the presence of two categories of precursor cells that reside within the organs and tissues of postnatal animals. These two categories of precursor cells are lineage-committed (multipotent, tripotent, bipotent, and unipotent) progenitor cells and lineage-uncommitted pluripotent (epiblastic-like, ectodermal, mesodermal, and endodermal) stem cells. These reserve precursor cells provide for the continual maintenance and repair of the organism after birth.
Asunto(s)
Células Madre Mesenquimatosas/citología , Células Madre Mesenquimatosas/fisiología , Células Madre Pluripotentes/citología , Células Madre Pluripotentes/fisiología , Adulto , Animales , Linaje de la Célula/fisiología , HumanosRESUMEN
Lineage uncommitted pluripotent stem cells reside in the connective tissue of skeletal muscle. The present study was carried out with pluripotent stem cells (PPSCs) isolated from 6-month old rat muscle. Before differentiation, these cells were vimentin+, CD90+, CD45-, and varied in their expression of CD34. The PPSCs were expanded as non-adherent aggregates under similar conditions to those used to generate neurospheres from embryonic or neural stem cells. The PPSC-derived neurospheres were positive for nestin, an early marker present in neuronal precursors, and expressed the two alternative mRNA forms of the neuroectodermal marker Pax-6, as well as mRNA for Oct-4, a gene related to the pluripotentiality of stem cells. To confirm their neural potential, PPSC-derived neurospheres were plated on coated coverslips under varying conditions: Neurobasal medium with N2 or B27, and either NT3 or BDNF. After 4-6 days the cells expressed neuronal (Tuj1+, NF68), astrocytic (GFAP) and oligodendrocytic (MOSP+, MBP+) markers, both by immunocytochemistry and RT-PCR. In addition, PPSCs were cultured as monolayers under adherent conditions, exposed to growth factors and defined differentiating conditions for 5 hr, and subsequently kept for 2 days in a maturation medium. At this point they gave rise to a mixed population of early neural progenitors (Nestin+ or NG2+), immature and mature neurons (Tuj1+ and NF145+) and myelin producing oligodendrocytes (CNPase + and MOSP+). Our study shows that PPSCs present in adult muscle can overcome germ lineage restrictions and express the molecular characteristics of brain cells. Therefore, PPSCs isolated from adult muscle could provide a novel source for autologous cell replacement in neurodegenerative and demyelinating diseases.