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Stem Cells: Relatively undifferentiated cells that retain the ability to divide and proliferate throughout postnatal life to provide progenitor cells that can differentiate into specialized cells.

Adult Stem Cells

JoVE 10810

Stem cells are undifferentiated cells that divide and produce more stem cells or progenitor cells that differentiate into mature, specialized cell types. All the cells in the body are generated from stem cells in the early embryo, but small populations of stem cells are also present in many adult tissues including the bone marrow, brain, skin, and gut. These adult stem cells typically produce the various cell types found in that tissue—to replace cells that are damaged or to continuously renew the tissue. The epithelium lining the small intestine is continuously renewed by adult stem cells. It is the most rapidly replaced tissue in the human body, with most cells being replaced within 3-5 days. The intestinal epithelium consists of thousands of villi that protrude into the interior of the small intestine—increasing its surface area to aid in the absorption of nutrients. Intestinal stem cells are located at the base of invaginations called crypts that lie between the villi. They divide to produce new stem cells, as well as daughter cells (called transit amplifying cells) that divide rapidly, move up the villi and differentiate into all the cell types in the intestinal epithelium, including absorptive, goblet, enteroendocrine, and Paneth cells. These mature cells continue to move up the villi as they carry out their functions, except Paneth cell

 Core: Biotechnology

Induced Pluripotent Stem Cells

JoVE 10812

Stem cells are undifferentiated cells that divide and produce different types of cells. Ordinarily, cells that have differentiated into a specific cell type are post-mitotic—that is, they no longer divide. However, scientists have found a way to reprogram these mature cells so that they “de-differentiate” and return to an unspecialized, proliferative state. These cells are also pluripotent like embryonic stem cells—able to produce all cell types—and are therefore called induced pluripotent stem cells (iPSCs). iPSCs are potentially valuable in medicine, because a patient who needs a particular cell type—for instance, someone with a damaged retina due to macular degeneration—could receive a transplant of the required cells, generated from another cell type in their own body. This is called autologous transplantation, and it reduces the risk of transplant rejection that can occur when tissues are transplanted between individuals. To create iPSCs, mature cells such as skin fibroblasts or blood cells from a person are grown in culture. Then, genes for multiple transcription factors are delivered into the cells using a viral vector, and the transcription factor proteins are expressed using the cell’s machinery. The transcription factors then turn on many other genes that are expressed by embryonic stem cells, re

 Core: Biotechnology

Tissue Regeneration with Somatic Stem Cells

JoVE 5339

Somatic or adult stem cells, like embryonic stem cells, are capable of self-renewal but demonstrate a restricted differentiation potential. Nonetheless, these cells are crucial to homeostatic processes and play an important role in tissue repair. By studying and manipulating this cell population, scientist may be able to develop new regenerative therapies for injuries and diseases.


 Developmental Biology

Embryonic Stem Cells

JoVE 10811

Embryonic stem (ES) cells are undifferentiated pluripotent cells, meaning they can produce any cell type in the body. This gives them tremendous potential in science and medicine since they can generate specific cell types for use in research or to replace body cells lost due to damage or disease.

ES cells are present in the inner cell mass of an embryo at the blastocyst stage, which occurs at about 3–5 days after fertilization in humans before the embryo is implanted in the uterus. Human ES cells are usually derived from donated embryos left over from the in vitro fertilization (IVF) process. The cells are collected and grown in culture, where they can divide indefinitely—creating ES cell lines. Under certain conditions, ES cells can differentiate—either spontaneously into a variety of cell types, or in a directed fashion to produce desired cell types. Scientists can control which cell types are generated by manipulating the culture conditions—such as changing the surface of the culture dish or adding specific growth factors to the culture medium—as well as by genetically modifying the cells. Through these methods, researchers have been able to generate many specific cell types from ES cells, including blood, nerve, heart, bone, liver, and pancreas cells. Regenerative medicine concerns the creation of living, functio

 Core: Biotechnology

An Introduction to Stem Cell Biology

JoVE 5331

Cells that can differentiate into a variety of cell types, known as stem cells, are at the center of one of the most exciting fields of science today. Stem cell biologists are working to understand the basic mechanisms that regulate how these cells function. These researchers are also interested in harnessing the remarkable potential of stem cells to treat human diseases.


Here,…

 Developmental Biology

Embryonic Stem Cell Culture and Differentiation

JoVE 5332

Culturing embryonic stem (ES) cells requires conditions that maintain these cells in an undifferentiated state to preserve their capacity for self-renewal and pluripotency. Stem cell biologists are continuously optimizing methods to improve the efficiency of ES cell culture, and are simultaneously trying to direct the differentiation of ES cells into specific cell types that could be used in…

 Developmental Biology

Passaging Cells

JoVE 5052

Cell lines are frequently used in biomedical experiments, as they allow rapid culture and expansion of cell types for experimental analysis. Cell lines are cultured under similar conditions when compared to freshly-isolated, or primary, cells, but with some basic important differences: (i) cell lines require their own specific growth factor cocktails and (ii) their growth must be more closely…

 Basic Methods in Cellular and Molecular Biology

Spermatogenesis

JoVE 10905

Spermatogenesis is the process by which haploid sperm cells are produced in the male testes. It starts with stem cells located close to the outer rim of seminiferous tubules. These spermatogonial stem cells divide asymmetrically to give rise to additional stem cells (meaning that these structures “self-renew”), as well as sperm progenitors, called spermatocytes. Importantly, this method of asymmetric mitotic division maintains a population of spermatogonial stem cells in the male reproductive tract, ensuring that sperm will continue to be produced throughout a man’s lifespan. As spermatogenesis proceeds, spermatocytes embark on meiosis, and each ultimately divides to form four sperm—each with only 23 chromosomes— that are expelled into the male reproductive tract. Interestingly, this is in contrast to oogenesis in women, during which only a single egg is generated for every progenitor cell. At the end of spermatogenesis, sperm demonstrate their characteristic shape: a “head” harboring minimal cytoplasm and a highly condensed nucleus, as well as a motile tail (flagellum). They are small cells, with no organelles such as ribosomes, ER or Golgi, but do have many mitochondria around the flagellum for power. Just below the head is the acrosomal vesicle which contains hydrolytic enzymes to penetrate the egg outer coat—th

 Core: Reproduction and Development

Induced Pluripotency

JoVE 5333

Induced pluripotent stem cells (iPSCs) are somatic cells that have been genetically reprogrammed to form undifferentiated stem cells. Like embryonic stem cells, iPSCs can be grown in culture conditions that promote differentiation into different cell types. Thus, iPSCs may provide a potentially unlimited source of any human cell type, which is a major breakthrough in the field of regenerative…

 Developmental Biology

Inflammation

JoVE 10902

In response to tissue injury and infection, mast cells initiate inflammation. Mast cells release chemicals that increase the permeability of adjacent blood capillaries and attract additional immune cells to the wound or site of infection. Neutrophils are phagocytic leukocytes that exit the bloodstream and engulf invading microbes. Blood clotting platelets seal the wound and fibers create a scaffold for wound healing. Macrophages engulf aging neutrophils to end the acute inflammatory response. Tissue injury and infection are the primary causes of acute inflammation. Inflammation protects the body by eliminating the cause of tissue injury and initiating the removal of cell debris resulting from the initial damage and related immune cell activity. Inflammation involves mediators of both the innate and adaptive immune system. Proper regulation of inflammation is crucial to clear the pathogen and remove cell debris without overly damaging healthy tissue in the process. If inflammatory processes are not properly regulated, chronic inflammation can arise that is often fatal. Mast cells are the first to respond to tissue injury, as they are primarily located in areas that have contact with the exterior: the skin, gut, and airways. Mast cells have an arsenal of receptors on their cell surface and can hence be activated by a wide variety of stimuli, such as mi

 Core: Immune System

An Introduction to Aging and Regeneration

JoVE 5337

Tissues are maintained through a balance of cellular aging and regeneration. Aging refers to the gradual loss of cellular function, and regeneration is the repair of damaged tissue generally mediated by preexisting adult or somatic stem cells. Scientists are interested in understanding the biological mechanisms behind these two complex processes. By doing so, researchers may be able to use…

 Developmental Biology

In-vitro Mutagenesis

JoVE 10813

To learn more about the function of a gene, researchers can observe what happens when the gene is inactivated or “knocked out,” by creating genetically engineered knockout animals. Knockout mice have been particularly useful as models for human diseases such as cancer, Parkinson’s disease, and diabetes.

Genes can be randomly knocked out, or specific genes can be targeted. To knock out a particular gene, an engineered piece of DNA called a targeting vector is used to replace the normal gene, thereby inactivating it. Targeting vectors have sequences on each end that are identical—or homologous— to the sequences flanking each side of the gene of interest. These homologous sequences allow the targeting vector to replace the gene through homologous recombination—a process that occurs naturally between DNA with similar sequences during meiosis. The targeting vector is introduced into mouse embryonic stem cells in culture, using methods such as electroporation—use of electric pulses to temporarily create pores in the cell membrane. Typically, to identify cells where the vector has properly replaced the gene, it is designed to include a positive selection marker—such as the gene for neomycin resistance (NeoR)—between the homologous regions; and a negative selection marker—such as th

 Core: Biotechnology

An Introduction to Developmental Neurobiology

JoVE 5207

Developmental neuroscience is a field that explores how the nervous system is formed, from early embryonic stages through adulthood. Although it is known that neural progenitor cells follow predictable stages of proliferation, differentiation, migration, and maturation, the mechanisms controlling the progression through each stage are incompletely understood. Studying…

 Neuroscience

Outcomes of Glycolysis

JoVE 11006

Nearly all the energy used by cells comes from the bonds that make up complex, organic compounds. These organic compounds are broken down into simpler molecules, such as glucose. Subsequently, cells extract energy from glucose over many chemical reactions—a process called cellular respiration.

Cellular respiration can take place in the presence or absence of oxygen, referred to as aerobic and anaerobic respiration, respectively. In the presence of oxygen, cellular respiration starts with glycolysis and continues with pyruvate oxidation, the citric acid cycle, and oxidative phosphorylation. Both aerobic and anaerobic cellular respiration start with glycolysis. Glycolysis yields a net gain of two pyruvate molecules, two NADH molecules, and two ATP molecules (four produced minus two used during energy-requiring glycolysis). In addition to these major products, glycolysis generates two water molecules and two hydrogen ions. In cells that carry out anaerobic respiration, glycolysis is the primary source of ATP. These cells use fermentation to convert NADH from glycolysis back into NAD+, which is required to continue glycolysis. Glycolysis is also the primary source of ATP for mature mammalian red blood cells, which lack mitochondria. Cancer cells and stem cells rely on aerobic glycolysis for ATP. Cells that use aerobic respiration cont

 Core: Cellular Respiration

Bone Structure

JoVE 10864

Within the skeletal system, the structure of a bone, or osseous tissue, can be exemplified in a long bone, like the femur, where there are two types of osseous tissue: cortical and cancellous.

Covering the cortical, or compact bone, is a membrane called the periosteum, which contains connective tissue, capillaries, and nerves. The outer, solid layer—found along the diaphysis, the shaft—forms a dense protective shell around the medullary canal—the cavity that stores yellow bone marrow, composed primarily of fat cells. This space is also covered in a thin lining—the endosteum in which bone growth, remodeling, and repair occur. Within the dense layer of cortical bone are osteons—structural units, arranged in concentric rings called lamellae, that contain osteoblasts—cells critical for bone formation and growth. These cells eventually mature into osteocytes in the hollow space, the lacuna. Through the center of each osteon runs the Haversian canal, which contains more blood and lymphatic vessels, as well as nerve fibers. Towards the rounded ends of the long bone, the epiphyses is the second type of osseous tissue, known as the cancellous, or spongy, bone. This inner layer is composed of a honeycomb-like network of trabeculae—grouped arrangements that form along the lines of stress points to maximize strengt

 Core: Musculoskeletal System

Transcription

JoVE 10794

Transcription is the process of synthesizing RNA from a DNA sequence by RNA polymerase. It is the first step in producing a protein from a gene sequence. Additionally, many other proteins and regulatory sequences are involved in the proper synthesis of messenger RNA (mRNA). Regulation of transcription is responsible for the differentiation of all the different types of cells and often for the proper cellular response to environmental signals. In eukaryotes, the DNA is first transcribed into a primary RNA, or pre-mRNA, that can be further processed into a mature mRNA to serve as a template for the synthesis of proteins. In prokaryotes such as bacteria, however, translation of RNA into polypeptides can begin while the transcription is still ongoing, as RNA can be quickly degraded. Transcription can also produce different kinds RNA molecules that do not code for protein, such as microRNAs, transfer RNA (tRNA), and ribosomal RNA (rRNA)—all of which contribute to protein synthesis. With few exceptions, all of the cells in the human body have the same genetic information in them, from neurons in the brain to muscle cells in the heart. So how do cells assume such diverse forms and functions? To a large extent, the answer lies in the regulation of transcription during development of the organism. Specifically, transcriptional regulation plays a central ro

 Core: DNA Structure and Function

Tissues

JoVE 10696

Cells with similar structure and function are grouped into tissues. A group of tissues with a specialized function is called an organ. There are four main types of tissue in vertebrates: epithelial, connective, muscle, and nervous.

Epithelial tissue consists of thin sheets of cells and includes the skin and the linings of internal organs and body cavities. Epithelial cells are tightly packed, providing a barrier against injury, infection, and water loss. Epithelial tissue can be a single layer called simple epithelium, or multiple layers called stratified epithelium. In stratified epithelium, such as the skin, the outer cells—which are subject to damage—are replaced through the division of cells underneath. Epithelial cells have a variety of shapes, including squamous (flattened), cuboid, and columnar. Some epithelial tissues absorb or secrete substances, such as the lining of the intestines. Connective tissue is composed of cells within an extracellular matrix and includes loose connective tissue, fibrous connective tissue, adipose (fat) tissue, cartilage, bone, and blood. Although the characteristics of connective tissue vary greatly, their general function is to support and attach multiple tissues. For example, tendons are made of fibrous connective tissue and attach muscle to bone. Blood transports oxygen, nutrients and waste produ

 Core: Cell Structure and Function

Replication in Eukaryotes

JoVE 10789

In eukaryotic cells, DNA replication is highly conserved and tightly regulated. Multiple linear chromosomes must be duplicated with high fidelity before cell division, so there are many proteins that fill specialized roles in the replication process. Replication occurs in three phases: initiation, elongation, and termination, and ends with two complete sets of chromosomes in the nucleus.

Eukaryotic replication follows many of the same principles as prokaryotic DNA replication, but because the genome is much larger and the chromosomes are linear rather than circular, the process requires more proteins and has a few key differences. Replication occurs simultaneously at multiple origins of replication along each chromosome. Initiator proteins recognize and bind to the origin, recruiting helicase to unwind the DNA double helix. At each point of origin, two replication forks form. Primase then adds short RNA primers to the single strands of DNA, which serve as a starting point for DNA polymerase to bind and begin copying the sequence. DNA can only be synthesized in the 5’ to 3’ direction, so replication of both strands from a single replication fork proceeds in two different directions. The leading strand is synthesized continuously, while the lagging strand is synthesized in short stretches 100-200 base pairs in length, called Okazaki fragments. Once the bu

 Core: DNA Structure and Function

Cell Cycle Analysis

JoVE 5641

Cell cycle refers to the set of events through which a cell grows, replicates its genome, and ultimately divides into two daughter cells through the process of mitosis. Because the amount of DNA in a cell shows characteristic changes throughout the cycle, techniques known as cell cycle analysis can be used to separate a population of cells according to the different phases …

 Cell Biology

Recombineering and Gene Targeting

JoVE 5553

One of the most widely used tools in modern biology is molecular cloning with restriction enzymes, which create compatible ends between DNA fragments that allow them to be joined together. However, this technique has certain restrictions that limit its applicability for large or complex DNA construct generation. A newer technique that addresses some of these shortcomings…

 Genetics

Fate Mapping

JoVE 5335

Fate mapping is a technique used to understand how embryonic cells divide, differentiate, and migrate during development. In classic fate mapping experiments, cells in different areas of an embryo are labeled with a chemical dye and then tracked to determine which tissues or structures they form. Technological improvements now allow for individual cells to be marked and traced throughout…

 Developmental Biology

Histotypic Tissue Culture

JoVE 5787

Although two-dimensional tissue culture has been common for some time, cells behave more realistically in a three-dimensional culture, and more closely mimics native tissue. This video introduces histotypic tissue culture, where the growth and propagation of one cell line is done in an engineered three-dimensional matrix to reach high cell density. Here, we show the…

 Bioengineering

Overview of Tissue Engineering

JoVE 5785

Tissue engineering is an emerging field, which aims to create artificial tissue from biomaterials, specific cells and growth factors. These engineered tissue constructs have far-reaching benefits, with possibilities for organ replacement and tissue repair.


This video introduces the field of tissue engineering and examines the components …

 Bioengineering

Whole Organ Tissue Culture

JoVE 5799

Whole organs can be cultured ex vivo using specialized bioreactors, with the goal of repairing or replacing entire organs. This method uses a donor organ that is stripped of all cells, leaving behind the three-dimensional structure, and is then repopulated with new cells. This video demonstrates the whole organ culture of lungs, and shows how a dynamic culture…

 Bioengineering

Cleavage and Blastulation

JoVE 10908

After a large-single-celled zygote is produced via fertilization, the process of cleavage occurs while zygotes travel through the uterine tube. Cleavage is a mitotic cell division that does not result in growth. With each round of successive cell division, daughter cells get increasingly smaller.

At the beginning of embryogenesis, maternal mRNAs control development. However, by the eight-cell stage of cleavage, embryonic genes become activated in a process called zygotic genome activation (ZGA). As a result, maternal mRNAs get degraded, and ZGA causes a transition from maternal to zygotic genetic control of developing an embryo. Although maternal mRNAs get degraded, previously translated proteins may remain in the embryo through later stages of development. Cleavage patterns vary between organisms depending on the presence and distribution of egg yolk amongst other factors. For example, mammals have a holoblastic rotational cleavage pattern. They are holoblastic because they have sparse, but evenly distributed yolk and therefore end up with a cleavage furrow that extends through the entire embryo as opposed to being meroblastic where the cleavage furrow does not extend through the yolk-dense portion of the cytoplasm. At the onset of cleavage, rotational cleavage begins when the zygote first divides to form two smaller daughter cells called blas

 Core: Reproduction and Development

An Introduction to Organogenesis

JoVE 5334

Organogenesis is the process by which organs arise from one of three germ layers during the later stages of embryonic development. Researchers studying organogenesis want to better understand the genetic programs, cell-cell interactions, and mechanical forces involved in this process. Ultimately, scientists hope to use this knowledge to create therapies and artificial organs that will help…

 Developmental Biology

C. elegans Development and Reproduction

JoVE 5110

Ceanorhabditis elegans is a powerful tool to help understand how organisms develop from a single cell into a vast interconnected array of functioning tissues. Early work in C. elegans traced the complete cell lineage and structure at the electron microscopy level, allowing researchers unprecedented insight into the connection between genes, development and disease. …

 Biology I

Genome Editing

JoVE 5554

A well-established technique for modifying specific sequences in the genome is gene targeting by homologous recombination, but this method can be laborious and only works in certain organisms. Recent advances have led to the development of “genome editing”, which works by inducing double-strand breaks in DNA using engineered nuclease enzymes guided to target…

 Genetics

Development and Reproduction of the Laboratory Mouse

JoVE 5159

Successful breeding of the laboratory mouse (Mus musculus) is critical to the establishment and maintenance of a productive animal colony. Additionally, mouse embryos are frequently studied to answer questions about developmental processes. A wide variety of genetic tools now exist for regulating gene expression during mouse embryonic and postnatal development, which can help…

 Biology II

Genetic Engineering of Model Organisms

JoVE 5327

Transgenesis, or the use of genetic engineering to alter gene expression, is widely used in the field of developmental biology. Scientists use a number of approaches to alter the function of genes to understand their roles in developmental processes. This includes replacement of a gene with a nonfunctional copy, or adding a visualizable tag to a gene that allows the resultant fusion protein to …

 Developmental Biology

Blood Withdrawal I

JoVE 10246

Source: Kay Stewart, RVT, RLATG, CMAR; Valerie A. Schroeder, RVT, RLATG. University of Notre Dame, IN


Blood collection is a common requirement for research studies that involve mice and rats. The method of blood withdrawal in mice and rats is dependent upon the volume of blood needed, the frequency of the sampling, the health status of the …

 Lab Animal Research

Bone Remodeling

JoVE 10866

Bone remodeling is a continuous and balanced process of bone resorption by osteoclasts and bone formation by osteoblasts. In adults, it helps maintain bone mass and calcium homeostasis. While mechanical stress can stimulate turnover as part of the normal maintenance and reparative process, several hormones also regulate bone remodeling.

Parathyroid hormone (PTH) maintains homeostatic control of blood calcium levels by regulating bone resorption. PTH is released from the parathyroid glands in response to low levels of calcium in the blood. It stimulates osteoblasts to produce immune molecules that promote the differentiation of precursor cells into osteoclasts. Activation of osteoclasts promotes bone resorption, causing the mineralized bone matrix to break down and release calcium into the blood. When blood calcium levels are restored, a negative-feedback loop prevents further release of PTH. Osteoporosis is a disease in which bone resorption exceeds bone formation, resulting in reduced bone density. Osteoporosis is more prevalent in women, especially after menopause. This is due to the critical role played by the female sex hormone—estrogen—in bone remodeling. Estrogen limits the formation of osteoclasts and promotes their destruction via apoptosis. This ensures that bone formation is higher than bone resorption. However, estrogen levels decli

 Core: Musculoskeletal System

Drosophila Larval IHC

JoVE 5106

Immunohistochemistry (IHC) is a technique used to visualize the presence and location of proteins within tissues. Drosophila larvae are particularly amenable to IHC because of the ease with which they can be processed for staining. Additionally, the larvae are transparent, meaning that some tissues can be visualized without the need for dissection.


In IHC, proteins are…

 Biology I

Transplantation Studies

JoVE 5336

Many developmental biologists are interested in the molecular signals and cellular interactions that induce a group of cells to develop into a particular tissue. To investigate this, scientists can use a classic technique known as transplantation, which involves tissue from a donor embryo being excised and grafted into a host embryo. By observing how transplanted tissues develop in host…

 Developmental Biology

Sterile Tissue Harvest

JoVE 10298

Source: Kay Stewart, RVT, RLATG, CMAR; Valerie A. Schroeder, RVT, RLATG. University of Notre Dame, IN


In 1959 The 3 R's were introduced by W.M.S. Russell and R.L. Burch in their book The Principles of Humane Experimental Technique. The 3 R's are replacement, reduction, and refinement of the use of animals in research.1 The …

 Lab Animal Research

An Overview of Gene Expression

JoVE 5546

Gene expression is the complex process where a cell uses its genetic information to make functional products. This process is regulated at multiple stages, and any misregulation could lead to diseases such as cancer.

This video highlights important historical discoveries relating to gene expression, including the…

 Genetics

Compound Administration I

JoVE 10198

Source: Kay Stewart, RVT, RLATG, CMAR; Valerie A. Schroeder, RVT, RLATG. University of Notre Dame, IN


As many research protocols require that a substance be injected into an animal, the route of delivery and the amount of the substance must be accurately determined. There are several routes of administration available in the mouse and rat. …

 Lab Animal Research

Evaluating the Heat Transfer of a Spin-and-Chill

JoVE 10440

Source: Michael G. Benton and Kerry M. Dooley, Department of Chemical Engineering, Louisiana State University, Baton Rouge, LA


The Spin-and-Chill uses heat transfer and fluid flow fundamentals to chill beverages from room temperature to 38 °F in as little as 2 min. It would take a refrigerator approximately 240 min and an ice chest…

 Chemical Engineering

Generation of Integration-free Induced Pluripotent Stem Cells from Human Peripheral Blood Mononuclear Cells Using Episomal Vectors

1State Key Laboratory of Experimental Hematology, Institute of Hematology and Blood Disease Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, 2Division of Regenerative Medicine, Department of Medicine, Loma Linda University, 3Department of Orthopaedic Surgery, Loma Linda University, 4Center for Stem Cell Medicine, Chinese Academy of Medical Sciences, 5Department of Stem Cell & Regenerative Medicine, Peking Union Medical College, 6Collaborative Innovation Center for Cancer Medicine, 7Tianjin Key Laboratory of Blood Cell Therapy and Technology

JoVE 55091

 Developmental Biology
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