Manipulation of stem cells and application in medicine pdf
File Name: manipulation of stem cells and application in medicine .zip
- Stem Cell Research
- Stem Cells Applications in Regenerative Medicine and Disease Therapeutics
- Mesenchymal stem cell perspective: cell biology to clinical progress
Stem Cell Research
Embryonic stem cells ES cells or ESCs are pluripotent stem cells derived from the inner cell mass of a blastocyst , an early-stage pre- implantation embryo. Isolating the embryoblast , or inner cell mass ICM results in destruction of the blastocyst, a process which raises ethical issues , including whether or not embryos at the pre-implantation stage should have the same moral considerations as embryos in the post-implantation stage of development. Researchers are currently focusing heavily on the therapeutic potential of embryonic stem cells, with clinical use being the goal for many laboratories.
However, adverse effects in the research and clinical processes such as tumours and unwanted immune responses have also been reported. Embryonic stem cells ESCs , derived from the blastocyst stage of early mammalian embryos, are distinguished by their ability to differentiate into any embryonic cell type and by their ability to self-renew. It is these traits that makes them valuable in the scientific and medical fields. ESCs have a normal karyotype , maintain high telomerase activity, and exhibit remarkable long-term proliferative potential.
Embryonic stem cells of the inner cell mass are pluripotent , meaning they are able to differentiate to generate primitive ectoderm, which ultimately differentiates during gastrulation into all derivatives of the three primary germ layers : ectoderm , endoderm , and mesoderm.
These germ layers generate each of the more than cell types in the adult human body. When provided with the appropriate signals, ESCs initially form precursor cells that in subsequently differentiate into the desired cell types.
Pluripotency distinguishes embryonic stem cells from adult stem cells , which are multipotent and can only produce a limited number of cell types. Under defined conditions, embryonic stem cells are capable of self-renewing indefinitely in an undifferentiated state. Self-renewal conditions must prevent the cells from clumping and maintain an environment that supports an unspecialized state.
ESCs divide very frequently due to a shortened G1 phase in their cell cycle. Rapid cell division allows the cells to quickly grow in number, but not size, which is important for early embryo development.
Although the shortened G1 phase has been linked to maintenance of pluripotency,  ESCs grown in serum-free 2i conditions do express hypo-phosphorylated active Retinoblastoma proteins and have an elongated G1 phase. Due to their plasticity and potentially unlimited capacity for self-renewal, embryonic stem cell therapies have been proposed for regenerative medicine and tissue replacement after injury or disease. Pluripotent stem cells have shown promise in treating a number of varying conditions, including but not limited to: spinal cord injuries , age related macular degeneration , diabetes , neurodegenerative disorders such as Parkinson's disease , AIDS , etc.
There are some ethical controversies surrounding this though see Ethical debate section below. Aside from these uses, ESCs can also be used for research on early human development, certain genetic disease, and in vitro toxicology testing.
According to a article in PNAS , "Human embryonic stem cells have the potential to differentiate into various cell types, and, thus, may be useful as a source of cells for transplantation or tissue engineering. Current research focuses on differentiating ESCs into a variety of cell types for eventual use as cell replacement therapies CRTs. Some of the cell types that have or are currently being developed include cardiomyocytes CM , neurons , hepatocytes , bone marrow cells, islet cells and endothelial cells.
For example, studies are underway to differentiate ESCs into tissue specific CMs and to eradicate their immature properties that distinguish them from adult CMs. Besides becoming an important alternative to organ transplants, ESCs are also being used in field of toxicology and as cellular screens to uncover new chemical entities NCEs that can be developed as small molecule drugs. Studies have shown that cardiomyocytes derived from ESCs are validated in vitro models to test drug responses and predict toxicity profiles.
ESC-derived hepatocytes are also useful models that could be used in the preclinical stages of drug discovery. However, the development of hepatocytes from ESCs has proven to be challenging and this hinders the ability to test drug metabolism.
Therefore, current research is focusing on establishing fully functional ESC-derived hepatocytes with stable phase I and II enzyme activity. Several new studies have started to address the concept of modeling genetic disorders with embryonic stem cells. Either by genetically manipulating the cells, or more recently, by deriving diseased cell lines identified by prenatal genetic diagnosis PGD , modeling genetic disorders is something that has been accomplished with stem cells.
This approach may very well prove valuable at studying disorders such as Fragile-X syndrome , Cystic fibrosis , and other genetic maladies that have no reliable model system. Yury Verlinsky , a Russian-American medical researcher who specialized in embryo and cellular genetics genetic cytology , developed prenatal diagnosis testing methods to determine genetic and chromosomal disorders a month and a half earlier than standard amniocentesis.
The techniques are now used by many pregnant women and prospective parents, especially couples who have a history of genetic abnormalities or where the woman is over the age of 35 when the risk of genetically related disorders is higher.
In addition, by allowing parents to select an embryo without genetic disorders, they have the potential of saving the lives of siblings that already had similar disorders and diseases using cells from the disease free offspring. For instance, human foreskin fibroblasts, one type of somatic cell, use non-homologous end joining NHEJ , an error prone DNA repair process, as the primary pathway for repairing double-strand breaks DSBs during all cell cycle stages.
ES cells use a different strategy to deal with DSBs. Consequently, robust mechanisms are needed in ES cells to repair DNA damages accurately, and if repair fails, to remove those cells with un-repaired DNA damages.
HRR can accurately repair DSBs in one sister chromosome by using intact information from the other sister chromosome. Cells in the G1 phase of the cell cycle i. On January 23, , Phase I clinical trials for transplantation of oligodendrocytes a cell type of the brain and spinal cord derived from human ES cells into spinal cord-injured individuals received approval from the U.
West , PhD. A previous experiment had shown an improvement in locomotor recovery in spinal cord-injured rats after a 7-day delayed transplantation of human ES cells that had been pushed into an oligodendrocytic lineage. The researchers emphasized that the injections were not expected to fully cure the patients and restore all mobility.
Based on the results of the rodent trials, researchers speculated that restoration of myelin sheathes and an increase in mobility might occur. This first trial was primarily designed to test the safety of these procedures and if everything went well, it was hoped that it would lead to future studies that involve people with more severe disabilities. In November Geron announced it was halting the trial and dropping out of stem cell research for financial reasons, but would continue to monitor existing patients, and was attempting to find a partner that could continue their research.
Michael D. West , acquired all of Geron's stem cell assets, with the stated intention of restarting Geron's embryonic stem cell-based clinical trial for spinal cord injury research.
Supported by California public funds, CIRM is the largest funder of stem cell-related research and development in the world. The award provides funding for Asterias to reinitiate clinical development of AST-OPC1 in subjects with spinal cord injury and to expand clinical testing of escalating doses in the target population intended for future pivotal trials.
OPCs and their mature derivatives called oligodendrocytes provide critical functional support for nerve cells in the spinal cord and brain.
Asterias recently presented the results from phase 1 clinical trial testing of a low dose of AST-OPC1 in patients with neurologically-complete thoracic spinal cord injury. Patients followed 2—3 years after AST-OPC1 administration showed no evidence of serious adverse events associated with the cells in detailed follow-up assessments including frequent neurological exams and MRIs.
Immune monitoring of subjects through one year post-transplantation showed no evidence of antibody-based or cellular immune responses to AST-OPC1. In four of the five subjects, serial MRI scans performed throughout the 2—3 year follow-up period indicate that reduced spinal cord cavitation may have occurred and that AST-OPC1 may have had some positive effects in reducing spinal cord tissue deterioration.
The major concern with the possible transplantation of ESC into patients as therapies is their ability to form tumors including teratoma. The main strategy to enhance the safety of ESC for potential clinical use is to differentiate the ESC into specific cell types e.
Following differentiation, the cells are subjected to sorting by flow cytometry for further purification. ESC are predicted to be inherently safer than IPS cells created with genetically-integrating viral vectors because they are not genetically modified with genes such as c-Myc that are linked to cancer.
Nonetheless, ESC express very high levels of the iPS inducing genes and these genes including Myc are essential for ESC self-renewal and pluripotency,  and potential strategies to improve safety by eliminating c-Myc expression are unlikely to preserve the cells' "stemness". However, N-myc and L-myc have been identified to induce iPS cells instead of c-myc with similar efficiency.
Due to the nature of embryonic stem cell research, there are a lot of controversial opinions on the topic. Since harvesting embryonic stem cells necessitates destroying the embryo from which those cells are obtained, the moral status of the embryo comes into question.
Some people claim that the 5-day old mass of cells is too young to achieve personhood or that the embryo, if donated from an IVF clinic which is where labs typically acquire embryos from , would otherwise go to medical waste anyway. Opponents of ESC research claim that an embryo is a human life, therefore destroying it is murder and the embryo must be protected under the same ethical view as a more developed human being.
In vitro fertilization generates multiple embryos. The surplus of embryos is not clinically used or is unsuitable for implantation into the patient, and therefore may be donated by the donor with consent. Human embryonic stem cells can be derived from these donated embryos or additionally they can also be extracted from cloned embryos using a cell from a patient and a donated egg. Immunosurgery, the process in which antibodies are bound to the trophectoderm and removed by another solution, and mechanical dissection are performed to achieve separation.
The resulting inner cell mass cells are plated onto cells that will supply support. The inner cell mass cells attach and expand further to form a human embryonic cell line, which are undifferentiated. These cells are fed daily and are enzymatically or mechanically separated every four to seven days. For differentiation to occur, the human embryonic stem cell line is removed from the supporting cells to form embryoid bodies, is co-cultured with a serum containing necessary signals, or is grafted in a three-dimensional scaffold to result.
Embryonic stem cells are derived from the inner cell mass of the early embryo , which are harvested from the donor mother animal. Martin Evans and Matthew Kaufman reported a technique that delays embryo implantation, allowing the inner cell mass to increase. This process includes removing the donor mother's ovaries and dosing her with progesterone , changing the hormone environment, which causes the embryos to remain free in the uterus.
Clonal cell lines are created by growing up a single cell. Evans and Kaufman showed that the cells grown out from these cultures could form teratomas and embryoid bodies , and differentiate in vitro, all of which indicating that the cells are pluripotent.
Gail Martin derived and cultured her ES cells differently. She removed the embryos from the donor mother at approximately 76 hours after copulation and cultured them overnight in a medium containing serum. The following day, she removed the inner cell mass from the late blastocyst using microsurgery.
The extracted inner cell mass was cultured on fibroblasts treated with mitomycin-c in a medium containing serum and conditioned by ES cells. After approximately one week, colonies of cells grew out. These cells grew in culture and demonstrated pluripotent characteristics, as demonstrated by the ability to form teratomas , differentiate in vitro, and form embryoid bodies. Martin referred to these cells as ES cells.
It is now known that the feeder cells provide leukemia inhibitory factor LIF and serum provides bone morphogenetic proteins BMPs that are necessary to prevent ES cells from differentiating. Furthermore, it has been demonstrated that different mouse strains have different efficiencies for isolating ES cells.
For human treatment, there is a need for patient specific pluripotent cells. Generation of human ES cells is more difficult and faces ethical issues. So, in addition to human ES cell research, many groups are focused on the generation of induced pluripotent stem cells iPS cells.
On August 23, , the online edition of Nature scientific journal published a letter by Dr. Robert Lanza medical director of Advanced Cell Technology in Worcester, MA stating that his team had found a way to extract embryonic stem cells without destroying the actual embryo.
In March, , the limitation was lifted. Human embryonic stem cells have also been derived by somatic cell nuclear transfer SCNT. However, in this case SCNT was used to produce embryonic stem cell lines in a lab, not living organisms via a pregnancy.
The "therapeutic" part of the name is included because of the hope that SCNT produced embryonic stem cells could have clinical utility. In it was shown that pluripotent stem cells highly similar to embryonic stem cells can be generated by the delivery of three genes Oct4 , Sox2 , and Klf4 to differentiated cells.
Because ethical concerns regarding embryonic stem cells typically are about their derivation from terminated embryos, it is believed that reprogramming to these "induced pluripotent stem cells" iPS cells may be less controversial.
Stem Cells Applications in Regenerative Medicine and Disease Therapeutics
Metrics details. Stem cells are a relevant source of information about cellular differentiation, molecular processes and tissue homeostasis, but also one of the most putative biological tools to treat degenerative diseases. This review focuses on human stem cells clinical and experimental applications. Our aim is to take a correct view of the available stem cell subtypes and their rational use in the medical area, with a specific focus on their therapeutic benefits and side effects. We have reviewed the main clinical trials dividing them basing on their clinical applications, and taking into account the ethical issue associated with the stem cell therapy.
Review Series Free access Phone: ; Fax: ; E-mail: insoo. Find articles by Hyun, I. Published January 4, - More info. Discussion of the bioethics of human stem cell research has transitioned from controversies over the source of human embryonic stem cells to concerns about the ethical use of stem cells in basic and clinical research.
Mesenchymal stem cell perspective: cell biology to clinical progress
Regenerative Medicine and Tissue Engineering. Advances in stem cell research and media publicity of stem cell potential have raised the hopes of patients with severe disabilities and conditions which lack a cure. While stem-cell-based therapies are the clinical standard of care for a few conditions, such as leukemia and more recently for some burns and corneal disorders, stem cell tourism continues to rise worldwide.
Most cells in the body are differentiated cells. These cells can only serve a specific purpose in a particular organ. For example, red blood cells are specifically designed to carry oxygen through the blood. All humans start out as only one cell.
Not a MyNAP member yet? Register for a free account to start saving and receiving special member only perks. S tem cell research offers unprecedented opportunities for developing new medical therapies for debilitating diseases and a new way to explore fundamental questions of biology.
Создатель последнего шифра, который никто никогда не взломает. Сьюзан долго молчала. - Но… это значит… Стратмор посмотрел ей прямо в глаза: - Да. Энсей Танкадо только что превратил ТРАНСТЕКСТ в устаревшую рухлядь. ГЛАВА 6 Хотя Энсей Танкадо еще не родился, когда шла Вторая мировая война, он тщательно изучал все, что было о ней написано, - особенно о кульминации войны, атомном взрыве, в огне которого сгорело сто тысяч его соотечественников. Хиросима, 6 августа 1945 года, 8.
Мой Бог. Это была настоящая красотка.