查看更多>>摘要：Therapeutic cloning refers to the derivation of embryonic stem cells (ntESC) from embryos derived from somatic cell nuclear transfer (SCNT) also known as cloning. Cloning involves transplanting a differentiated cell into an oocyte that has had its nucleus (DNA) removed. The reconstructed oocyte can be activated to divide and develop into an embryo. The process that allows this to happen is termed nuclear reprogramming, and is defined as the mechanism through which a differentiated cell de-differentiates or returns to a totipotent state (capable of giving rise to any cell type, including extra-embryonic) and directs embryonic development. Cells from blastocyst stage cloned embryos can be used to generate ntESC lines.
查看更多>>摘要：Tuberous sclerosis complex (TSC) is an autosomal dominant neurocutaneous disorder characterized by the presence of multiple hamartomas in almost every organ, most notably in the skin, brain, heart, kidneys, liver, and lungs . The classic syndromes of TSC are seizures, mental retardation, and cutaneous angiofibromas . Two tumor suppressor genes, Tsc1 and Tsc2, have been identified for pathogenesis of TSC. Tscl is located on chromosome 9q34 and encodes for the protein hamartin (130 kDa). Tsc2 is located on chromosome 16pl3.3 and encodes for the protein tuberin (180 kDa).
查看更多>>摘要：Countries in North America and Europe have the highest incidence of inflammatory disorders of the gastrointestinal tract. The prevalence of inflammatory bowel diseases (IBD), which comprise Crohn's disease (CD) and ulcerative colitis (UC), now ranges from 10-200 cases per 100 000 individuals . Although new therapeutic approaches have been developed to improve current treatments, the etiology of this disorder remains elusive. Crohn's disease and ulcerative colitis are known to show similar clinical and pathological characteristics; however it is now believed that these two forms of IBD are entirely different.
查看更多>>摘要：Most defective and unwanted cells die by apoptosis, an exquisitely controlled genetic programme for removing such cells without damaging the surrounding tissue. Once a cell has committed to apoptosis, the process is remarkably efficient, and is completed within a few minutes of initiation. This point of no return for an apoptotic cell is commonly held to be the point at which the outer mitochondrial membrane is permeabilised, a process regulated by the Bcl-2 family of proteins. How these proteins regulate this decision point is central to diseases such as cancer where apoptotic control is lost. In this review, we will discuss apoptotic signalling and how a cell makes the irreversible decision to die. We will focus on one set of survival signals, those derived by cell adhesion to the extracellular matrix (ECM), and use these to highlight the complexities of apoptotic signalling. In particular, we will illustrate how multiple signalling pathways converge to determine critical cell fate decisions.
查看更多>>摘要：Numerous membrane proteins are cleaved by tumor necrosis factor-α converting enzyme (TACE), which causes the release of their ectodomains. An ADAM (a disintegrin and metalloprotease domain) family member, TACE contains several noncatalytic domains whose roles in ectodomain shedding have yet to be fully resolved. Here, we have explored the function of the transmembrane domain (TM) of TACE by coupling molecular engineering and functional analysis. A TM-free TACE construct that is anchored to the plasma membrane by a glycosylphosphatidylino-sitol (GPI)-binding polypeptide failed to restore shedding of transforming growth factor-α (TGF-α), tumor necrosis factor-α (TNF-α) and L-selectin in cells lacking endogenous TACE activity. Substitution of the TACE TM with that of the prolactin receptor or platelet-derived growth factor receptor (PDGFR) also resulted in severe loss of TGF-α shedding, but had no effects on the cleavage of TNF-α and L-selectin. Replacement of the TM in TGF-α with that of L-selectin enabled TGF-α shedding by the TACE mutants carrying the TM of prolactin receptor and PDGFR. Taken together, our observations suggest that anchorage of TACE to the lipid bilayer through a TM is required for efficient cleavage of a broad spectrum of substrates, and that the amino-acid sequence of TACE TM may play a role in regulatory specificity among TACE substrates.
查看更多>>摘要：Homozygous human embryonic stem cells (hESCs) are thought to be better cell sources for hESC banking because their human leukocyte antigen (HLA) haplotype would strongly increase the degree of matching for certain populations with relatively smaller cohorts of cell lines. Homozygous hESCs can be generated from parthenogenetic embryos, but only heterozygous hESCs have been established using the current strategy to artificially activate the oocyte without second polar body extrusion. Here we report the first successful derivation of a human homozygous ESC line (chHES-32) from a one-pronuclear oocyte following routine in vitro fertilization treatment. chHES-32 cells express common markers and genes with normal hESCs. They have been propagated in an undifferentiated state for more than a year ( > P50) and have maintained a stable karyotype of 46, XX. When differentiated in vivo and in vitro, chHES-32 cells can form derivatives from all three embryonic germ layers. The almost undetectable expression of five paternally expressed imprinted genes and their HLA genotype identical to the oocyte donor indicated their parthenogenetic origin. Using genome-wide single-nucleotide polymorphism analysis and DNA fingerprinting, the homozygosity of chHES-32 cells was further confirmed. The results indicated that 'unwanted' one-prbnuclear oocytes might be a potential source for human homozygous and parthenogenetic ESCs, and suggested an alternative strategy for obtaining homozygous hESC lines from parthenogenetic haploid oocytes.
查看更多>>摘要：Parthenogenesis is one of the main, and most useful, methods to derive embryonic stem cells (ESCs), which may be an important source of histocompatible cells and tissues for cell therapy. Here we describe the derivation and characterization of two ESC lines (hPES-1 and hPES-2) from in vitro developed blastocysts following parthenogenetic activation of human oocytes. Typical ESC morphology was seen, and the expression of ESC markers was as expected for alkaline phosphatase, octamer-binding transcription factor 4, stage-specific embryonic antigen 3, stage-specific embryonic antigen A, TRA-1-60, and TRA-1-81, and there was absence of expression of negative markers such as stage-specific embryonic antigen 1. Expression of genes specific for different embryonic germ layers was detected from the embryoid bodies (EBs) of both hESC lines, suggesting their differentiation potential in vitro. However, in vivo, only hPES-1 formed teratoma consisting of all three embryonic germ layers (hPES-2 did not). Interestingly, after continuous proliferation for more than 100 passages, hPES-1 cells still maintained a normal 46 XX karyotype; hPES-2 displayed abnormalities such as chromosome translocation after long term passages. Short Tandem Repeat (STR) results demonstrated that the hPES lines were genetic matches with the egg donors, and gene imprinting data confirmed the parthenogenetic origin of these ES cells. Genome-wide SNP analysis showed a pattern typical of parthenogenesis. All of these results demonstrated the feasibility to isolate and establish human parthenogenetic ESC lines, which provides an important tool for studying epigenetic effects in ESCs as well as for future therapeutic interventions in a clinical setting.
查看更多>>摘要：p28~(GANK) (also known as PSMD10, p28 and gankyrin) is an ankyrin repeat anti-apoptotic oncoprotein that is commonly overexpressed in hepatocellular carcinomas and increases the degradation of p53 and Rb. NF-κB (nuclear factor-κB) is known to be sequestered in the cytoplasm by IκB (inhibitor of NF-κB) proteins, but much less is known about the cytoplasmic retention of NF-κB by other cellular proteins. Here we show that p28~(GANK) inhibits NF-κB activity. As a nuclear-cytoplasmic shuttling protein, p28~(GANK) directly binds to NF-KB/RelA and exports RelA from nucleus through a chromosomal region maintenance-1 (CRM-1) dependent pathway, which results in the cytoplasmic retention of NF-κB/RelA. We demonstrate that all the ankyrin repeats of p28~(GANK) are required for the interaction with RelA and that the N terminus of p28~(GANK), which contains the nuclear export sequence (NES), is responsible for suppressing NF-κB/RelA nuclear translocation. These results suggest that overexpression of p28~(GANK) prevents the nuclear localization and inhibits the activity of NF-κB/RelA.
查看更多>>摘要：Sphingolipids have been suggested to act as second messengers for an array of cellular signaling activities in plant cells, including stress responses and programmed cell death (PCD). However, the mechanisms underpinning these processes are not well understood. Here, we report that an Arabidopsis mutant, fumonisin B1 resistant11-1 (fbr 11-1), which fails to generate reactive oxygen intermediates (ROIs), is incapable of initiating PCD when the mutant is challenged by fumonisin B_1 (FB_1), a specific inhibitor of ceramide synthase. Molecular analysis indicated that FBR11 encodes a long-chain basel (LCB1) subunit of serine palmitoyltransferase (SPT), which catalyzes the first rate-limiting step of de novo sphingolipid synthesis. Mass spectrometric analysis of the sphingolipid concentrations revealed that whereas the fbr 11-1 mutation did not affect basal levels of sphingoid bases, the mutant showed attenuated formation of sphingoid bases in response to FB_1. By a direct feeding experiment, we show that the free sphingoid bases dihydrosphingosine, phytosphingosine and sphingosine efficiently induce ROI generation followed by cell death. Conversely, ROI generation and cell death induced by dihydrosphingosine were specifically blocked by its phosphorylated form dihydrosphingosine-1-phosphate in a dose-dependent manner, suggesting that the maintenance of homeostasis between a free sphingoid base and its phosphorylated derivative is critical to determining the cell fate. Because alterations of the sphingolipid level occur prior to the ROI production, we propose that the free sphingoid bases are involved in the control of PCD in Arabidopsis, presumably through the regulation of the ROI level upon receiving different developmental or environmental cues.
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