Therapeutic cloning will in time allow scientists to create organs that are a perfect match for those in need of a transplant. The cloned organ would be based on the recipient’s genetic material and would not require the use of debilitating immunosuppressive therapies. There would also be no chance of rejection, which is fatal. Therapeutic cloning represents the ideal in organ transplantation, as it would provide an unlimited source of organs to anyone who needs them.
It is unfortunate that the term “cloning” refers to three very different procedures with three very different goals. It is also unfortunate that the first thought many people have when they hear the term is of horror movies which have showed the creation of human monsters or of armies of superhuman soldiers with subhuman brains. The reality of cloning is very different.
The three different types of “cloning” are:Embryo cloning: This is a medical technique which produces monozygotic (identical) twins or triplets. It duplicates the process that nature uses to produce twins or triplets. One or more cells are removed from a fertilized embryo and encouraged to develop into one or more duplicate embryos. Twins or triplets are thus formed, with identical DNA. This has been done for many years on various species of animals; only very limited experimentation has been done on humans.
Adult DNA cloning (a.k.a.  reproductive cloning) This technique which is intended to produce a duplicate of an existing animal. It has been used to clone a sheep and other mammals. The DNA from an ovum is removed and replaced with the DNA from a cell removed from an adult animal. Then, the fertilized ovum, now called a pre-embryo, is implanted in a womb and allowed to develop into a new animal. As of 2002-JAN, It had not been tried on humans. It is specifically forbidden by law in many countries. There are rumors that Dr. Severino Aninori has successfully initiated a pregnancy through reproductive cloning. It has the potential of producing a twin of an existing person. Based on previous animal studies, it also has the potential of producing severe genetic defects. For the latter reason alone, many medical ethicists consider it to be a profoundly immoral procedure when done on humans.
Therapeutic cloning (a.k.a. biomedical cloning): This is a procedure whose initial stages are identical to adult DNA cloning. However, the stem cells are removed from the pre-embryo with the intent of producing tissue or a whole organ for transplant back into the person  who supplied the DNA. The pre-embryo dies in the process.  The goal of therapeutic cloning is to produce a healthy copy of a sick person’s tissue or organ for transplant. This technique would be vastly superior to relying on organ transplants from other people. The supply would be unlimited, so there would be no waiting lists. The tissue or organ would have the sick person’s original DNA; the patient would not have to take immunosuppressant drugs for the rest of their life, as is now required after transplants. There would not be any danger of organ rejection.Author: dragon web profit

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Somatic cell nuclear transfer (SCNT) entails the removal of an oocyte nucleus followed by its replacement with a nucleus derived from a somatic cell obtained from that patient. Activation with chemicals or electric shock stimulates cell division up to the blastocyst stage at which time the inner cell mass is isolated and cultured, resulting in ESC. This approach is distinct from reproductive cloning because the blasotcyst is not transplanted back to the uterus. Hence, development does not proceed beyond the 100 cell stage. This process also differs from fertilization since no sperm is used in this process. The resulting ESC are perfectly matched to the patients immune system and no immunosuppressants would therefore be required to prevent rejection.
While interest in the field of nuclear cloning remains high since the birth of Dolly (1997), the first successful nuclear transfer was actually reported over fifty years ago by Briggs and King. Cloned frogs, which were the first vertebrates derived from nuclear transfer, were subsequently reported by Gurdon in 1962 although the nuclei were derived from non-adult sources. Indeed, in just the past six years alone important advances in nuclear cloning technology have been reported – a pace of discovery that betokens the relative immaturity of this research arena. In fact Dolly was not the first cloned mammal to be produced from adult cells. Live lambs were produced in 1996 using nuclear transfer and differentiated epithelial cells, although these were derived from embryonic discs. To be sure, the significance of the Dolly report was that this described the first mammal to be derived from an adult somatic cell using nuclear transfer. Subsequently, animals from several species have been grown using nuclear transfer technology, including cattle, goats, mice, and pigs.
A better understanding of the differences between reproductive cloning and therapeutic cloning may help alleviate some of the controversy surrounding these technologies. Banned in most countries for human applications, reproductive cloning is used to generate an embryo that has the identical genetic material as its cell source. Such an embryo could then be implanted into the uterus of a female to give rise to a liveborn infant that is a clone of the donor. In contrast, therapeutic cloning is used to generate only ESC lines whose genetic material is identical to that of its source. These autologous stem cells have the potential to become almost any type of cell in the adult body, and thus would be useful in tissue and organ replacement applications. Therefore, therapeutic cloning (SCNT) may provide an alternative source of transplantable cells. It has been estimated that approximately 3,000 people die every day in USA of diseases that could have been treated with stem cells-derived tissues. With current allogeneic tissue transplantation protocols, rejection is a frequent complication because of immunologic incompatibility and thus immunosuppressive drugs are generally required to manage host-versus-graft disease. The use of transplantable tissue and organs derived from therapeutic cloning could obviate unwanted immune responses typically associated with transplantation of non-autologous tissues.


While promising, somatic cell nuclear transfer technology has certain limitations requiring further improvement before it can be applied widely in clinical practice. Currently, the efficiency of the overall cloning process is quite low as the majority of embryos derived from animal cloning do not survive after implantation. In practical terms, multiple nuclear transfers must be performed in order to produce one live offspring for animal cloning applications. The potential for cloned embryos to grow into live offspring ranges between <1 and 18% for sheep, pigs, and mice. However, greater success (~ 80%) has been reported in cattle, a result which may in part be due to availability of advanced laboratory technologies specifically developed for this species for agricultural/breeding purposes. To improve cloning efficiencies, further improvements are required in the multiple complex steps of nuclear transfer such as enucleation and reconstruction, oocyte activation, and synchronization of cell cycle between donor cells and recipient oocytes.
It must be noted that abnormalities have been found in liveborn clones including macrosomia with an enlarged placenta ("large-offspring syndrome"), respiratory distress, defects of the kidney, liver, heart, and brain, obesity, and premature death. These may be related to epigenetics of cloned cells which involve reversible modifications of DNA, while the original DNA (genetic) sequences remain intact. Faulty epigenetic modulation in clones may result from altered DNA methylation and/or histone modifications causing the overall chromatin structure of somatic nuclei not to be reprogrammed to an embryonic pattern of expression. Reactivation of key embryonic genes at the blastocyst stage usually does not occur in embryos cloned from somatic cells, while embryos cloned from embryos consistently express early embryonic genes. Proper epigenetic reprogramming to an embryonic state may help to improve the cloning efficiency and reduce the incidence of abnormal cloned cells.
Novel applications of somatic cell nuclear transfer (therapeutic cloning)
We applied principles of both tissue engineering and therapeutic cloning in an effort to produce genetically identical renal tissue in an animal model (Bos taurus). Bovine skin fibroblasts from adult Holstein steers were obtained by ear notch and single donor cells were isolated and microinjected into the perivitelline space of donor enucleated oocytes (nuclear transfer). The resulting blastocysts were transferred to the uterus of progestin-synchronized recipients permit further in vivo growth. After 12 weeks cloned renal cells were harvested, expanded in vitro, then seeded onto biodegradable scaffolds. The constructs (consisting of cells + scaffolds) were then implanted into the subcutaneous space of the same steer from which the cells were cloned to allow for tissue growth.
The kidney is a complex organ with multiple cell types and a complex functional anatomy rendering it one of the most difficult organs to reconstruct. Previous efforts in tissue engineering of the kidney have been directed toward development of extracorporeal renal support systems made of biological and synthetic components. Although ex vivo renal replacement devices are known to be life-sustaining, there are obvious benefits for patients with end-stage kidney disease if such devices could be implanted long-term without the need for an extracorporeal perfusion circuit or immunosuppressive drugs.
Cloned renal cells were seeded on scaffolds consisting of three collagen-coated cylindrical polycarbonate membranes. The ends of the three membranes of each scaffold were connected to catheters terminating in a collecting reservoir. This created a renal neo-organ with a mechanism for collecting the excreted urinary fluid. Scaffolds with the collecting devices were transplanted subcutaneously into the same steer from which the genetic material originated and retrieved 12 weeks after implantation.


Renal unit seeded with cloned cells, three months after implantation, showing the accumulation of urinelike fluid. Chemical analysis of the urine-like fluid (for urea nitrogen/creatinine levels, electrolyte levels, specific gravity, and glucose concentration) revealed that the implanted renal cells possessed filtration, reabsorption, and secretory capabilities. Histological examination of the retrieved implants revealed extensive vascularization and self-organization of the cells into glomeruli- and tubule-like structures. A clear continuity between glomeruli, tubules, and the polycarbonate membrane was noted that allowed the passage of urine into the collecting reservoir. Immunohistochemical analysis with kidney-specific antibodies revealed the presence of renal proteins, and RT-PCR analysis confirmed the transcription of renal specific RNA in the cloned specimens. Western blot analysis confirmed the presence of elevated renal-specific protein levels.


As previous studies have confirmed bovine clones harbor mitochondrial DNA (mtDNA) of strictly oocyte origin, the donor egg’s mtDNA was thought to be a potential source of immunologic incompatibility. Differences in mtDNA-encoded proteins expressed by cloned cells could stimulate a T-cell response specific for mt-DNA-encoded minor histocompatibility antigens when cloned cells are implanted back into the original nuclear donor. We used nucleotide sequencing of the mtDNA genomes of the clone and fibroblast nuclear donor to identify potential antigens in the muscle constructs. Only two amino acid substitutions were noted to distinguish cells from the clone and the nuclear donor. Since peptide-binding motifs for bovine MHC class I molecules remain poorly understood, there is no reliable method to predict the impact of these amino acid substitutions on bovine histocompatibility.
Oocyte-derived mtDNA was also considered to be a potential source of immunologic incompatibility in cloned renal cells. Maternally transmitted minor histocompatibility antigens in mice have been shown to stimulate both skin allograft rejection in vivo and cytotoxic T lymphocytes expansion in vitro that could prevent the use of these cloned constructs in patients with chronic rejection of major histocompatibility-matched human renal transplants. We tested for a possible T-cell response to the cloned renal devices using delayed-type hypersensitivity testing in vivo and Elispot analysis of interferon-gamma secreting T-cells in vitro. Both analyses revealed that the cloned renal cells showed no evidence of T-cell response, suggesting that rejection will not necessarily occur in the presence of oocyte-derived mtDNA. This finding may represent a step forward in overcoming the histocompatibility problem of stem cell therapy.


These studies demonstrated that cells derived from nuclear transfer can be successfully harvested, expanded in culture, and transplanted in vivo with the use of biodegradable scaffolds on which the single suspended cells can organize into tissue structures that are genetically identical to that of the host. These studies were the first demonstration of the use of therapeutic cloning for regeneration of tissues in vivo. Others in the field have created mouse SCNT derived c-kit-positive stem cells to restore infarcted myocardium, dopaminergic neurons to correct the phenotype of a mouse model of Parkinson disease. The first HESC line derived from SCNT was created in February, 2004.
Author: dragon web profit

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Embryogenesis is a sequential process of differential gene expression dictated by the epigenetic environment. Exploiting epigenetic influence on phenotypic outcome, biotechnology platforms are developed for reversal of differentiation to achieve genetic reprogramming of adult sources back to an embryonic state. Such platforms include “therapeutic cloning” and “nuclear reprogramming” that bypass the need for embryo extraction to generate pluripotent stem cell phenotypes from autologous sources. Reprogramming of adult cells to generate customized embryonic-like stem cells offers the future for patient-specific regenerative therapies.
1. induced Pluripotent Stem Cells (iPSC) via nuclear reprogramming
Nuclear reprogramming of adults cells, namely skin fibroblasts, through ectopic introduction of a small number of pluripotency-associated transcription factors is an approach to induce an embryonic stem cell-like phenotype. In the mouse, such an approach has yielded induced pluripotent stem (iPS) cells sufficient for de novo embryogenesis and germline transmission. In humans, transcription factors associated with the pluripotency state were ranked through bioinformatic prioritization and further screened for reprogramming effectiveness through combinatorial viral delivery. Transcription factor sets Oct4, Sox2, c-Myc, and Klf4 or alternatively Oct4, Sox2, Nanog, and Lin28, are sufficient to reprogram human somatic cells to pluripotent stem cells that exhibit the essential characteristics of ESC, including maintenance of the developmental potential to differentiate into advanced derivatives of all three germ layers.
Moreover, iPS-based technology will facilitate the production of cell line panels that closely reflect the genetic diversity of a population enabling the discovery, development, and validation of therapies tailored for each individual. Alleviating technical limitations, including mutation through viral integration, incorporation of oncogenic genes, and establishing reliable patient-specific differentiation protocols are significant advancements that will facilitate clinical translation.
2. therapeutic cloning
Somatic cell nuclear transfer (SCNT) allows transacting factors present within the mammalian oocytes, conserved across species, to reprogram somatic cell nuclei to an undifferentiated state. Therapeutic cloning refers to SCNT in which the nuclear content of a somatic cell from an individual is transferred into an enucleated donor egg to derive blastocysts that contain pluripotent embryonic-like stem cells. In this way, SCNT has produced cloned ESCs from multiple mammalian somatic cell biopsies. The pluripotency of derived cells has been confirmed through germline transmission and reproductive cloning. However, due to technological limitations cloned human blastocysts have only recently been achieved although in low efficiency, and successful isolation of ESCs from the inner cell mass has yet to be demonstrated with human protocols.
Author: dragon web profit

Anti-Aging Skin Care Web Site: http://www.antiagingskincarebeauty.com

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