细胞凋亡研究进展（Research progress of apoptosis）

1、细胞凋亡研究进展（Research progress of apoptosis）Research progress of apoptosisAbstract:Cell apoptosis, oncosis and necrotic cell death is irreversible cell injury functional definition, apoptosis is one of the cell death, the total end changes and necrosis occurred in the living on behalf of local tissue af

2、ter cell death, is a kind of morphological diagnosis. In addition to cell apoptosis, there is another form of cell death or necrosis like death, said: the oncosis oncosis (oncosis), namely death (swelling death), was characterized by cell damage process of cytoplasmic swelling and karyolysis. Althou

3、gh there are still many problems and even the key questions are not clear, but the research in recent years the biochemical reaction mechanism in apoptosis signal transduction, cell apoptosis and apoptosis gene regulation and other aspects have made significant progress.Keywords: apoptosis, mitochon

4、drial cysteine proteaseIt is generally accepted that apoptosis plays an important role in cancer and other diseases. It has been widely studied in many biological fields. Since 1972, Kerr proposed the concept of cell apoptosis, from late 80s began to become a research hotspot in tumor etiology, path

5、ology, understanding of apoptosis gradually, the molecular mechanism of apoptosis occurred more and more thorough understanding, but also found that this process is not as simple as originally thought, but contains a complicated regulatory mechanism the.The stability of organism environment depends

6、not only on the proliferation and differentiation of cells, but also on the apoptosis of cells. Apoptosis (apoptosis), also known as programmed cell death, is one of the most important biological phenomena in biology. From nematodes to higher mammals, from embryos to adults, from physiology to patho

7、logy, from birth to death, different cells in the body have more of this form of death. More than 100 years ago, Carl Vogt had found this form of death. In 1972, Kerr et al. Re proposed the study and named apoptosis (apoptosis). Later, new insights have been made in the fields of morphology and bioc

8、hemistry of apoptosis.Recent studies have made clear progress in 3 areas. First, from the study of small nematode (C. elegans) apoptosis regulating gene, confirmed apoptosis is controlled by genetic development, among which Ced-9, Ced-3 and Ced-4 genes was determined, mammals also have corresponding

9、 homologous genes. Second, to elucidate the apoptotic signaling pathways, and to activate apoptotic cells by the activation of caspases (caspases), a specific death signaling molecule. Third, we study the relationship between apoptosis and some diseases, understand the pathogenesis of disease and ap

10、ply the theory and practice of apoptosis to prevent and cure diseases.Cloning of apoptosis related genesThe rapid development of apoptosis is due to the discovery of apoptosis related genes. In 1986, Professor Tsujimoto first cloned the gene Bcl-2 associated with type B lymphoma during his work in t

11、he United states. Later, it was proved that Bcl-2 gene plays a key role in regulating apoptosis. A large number of apoptotic genes have been cloned and their relationships have been studied from experimental animals elegans (Caenorhabditis). A total of 1090 cells are present in mature individuals of

12、 the nematode Caenorhabditis elegans, and 131 of these cells undergo apoptosis during development. The death of these cells is tightly regulated by genes. A number of nematode mutants have been isolated, and these mutants do not undergo apoptosis during the development of their apoptotic cells, resu

13、lting in developmental abnormalities. Professor Horvitz, of Harvard Medical School, first cloned apoptotic related genes from C.elegans and found that Ced-3 is a similar analogue of caspase, and Ced-4 is a novel apoptosis regulating protein.Importantly, scientists have found that Ced-9 is not only a

14、 nematode inhibitor of apoptosis but also a homologue of mammalian cell Bcl-2. This indicates that the mechanism of apoptosis is conserved in the evolutionary process. The study of apoptosis in C.elegans has established the basic framework of apoptosis. Know now,Ced-3 (mammalian cell caspase analog)

15、 and Ced-4 (APAF-1 analog in mammalian cells) are apoptotic promoting factors in c.elegans. Most other genes, such as Ced-1, Ced-2, Ced-5, Ced-6, Ced-7 and Ced-10, are responsible for phagocytosis and clearance of dead cell debris. Specifically, Ced-6 encodes a transporter called ABC (ATP-binding ca

16、ssette) that participates in the transport of cytoplasmic membrane proteins. Ced-7 participates in mutual recognition during phagocytosis and apoptosis. Ced-2, Ced-5 and Ced-10 may be involved in cell migration. Ced-5 encodes a DOCK180 (downstream, of, CRK, a proto oncogene protein) that is responsi

17、ble for stretching the surface of phagocytes to coat and engulf apoptotic cells. The molecular mechanisms involved in the phagocytosis of different phagocytic genes are not yet clear.Ced series of genes interact to modulate the apoptotic process. It can be said that the study of apoptosis during the

18、 development of C.elegans lays the foundation for the study of apoptosis in mammals and humans. Since then, scientists have cloned a large number of other animal genes related to apoptosis, that is, genes related to apoptosis related genes in c.elegans. Of course, the mechanism of apoptosis regulati

19、on in higher animals is more complex.With the study of the morphology of apoptotic cells, the changes of cellular biochemistry have been gradually elucidated. (1) calcium and protein kinase change: the accumulation and redistribution of intracellular Ca2+, the nucleus of Ca2+ increased 4, the activa

20、tion of endonuclease activity of protein kinase C in different cell types and different activity in a cycle. (2) endonuclease activation 5 a sequence of nucleotide fragments that are cut into 180 base pairs in DNA, showing a ladder like electrophoresis phenomenon. (3) to organize the accumulation an

21、d activation of transglutaminase. (4) the changes of cytoskeleton, such as actin. (5) the cell surface sugar chain, the phytohemagglutinin and the fibronectin receptor increase; the exposure of phosphatidylserine alters the biochemical characteristics of the cell surface.Two. Signal transduction of

22、apoptosisThe signal transduction pathways of apoptosis can be divided into two types: receptor mediated and non receptor mediated apoptotic pathways:Apoptosis mediated by 1. receptorsTumor necrosis factor ligand (TNF) and its receptor (TNFR) superfamily play a key role in immune regulation, and some

23、 members play an important role in the development of nervous system and other organs. At present, the cloned TNF ligands have 18, including TNF, FasL, TRAIL (TNF, related, apoptosis-inducing, ligand) and so on. These ligands are mostly transmembrane proteins, and their extracellular domains can be

24、cut by metalloproteinase and become soluble proteins. These molecules can form trimers or virus like polymers to play their biological effects. FasL is a type two transmembrane protein with a molecular weight of 40kD, i.e., the N terminal is in cytoplasm, and the C terminus of 150 amino acids is exp

25、osed to the extracellular region. The FasL and TNF superfamily ligands are 20% to 25% amino acids identical. FasL is expressed mainly in the eyes, anterior chamber, and testis and is expressed in CD8+Thl cells, CD4+T cells, and some NK cells. Its presence on the surface of the cell is very short, an

26、d the cleavage of the extracellular portion of the cell by metalloproteinases can become soluble and functional molecules.2. non receptor mediated apoptosis signal transductionCells maintain physiological activity and require various survival signals (surviving, signals). The loss and change of sign

27、als and extracellular matrix between adjacent cells, growth factors such as cytokines that support cell growth, and many kinds of viruses, a variety of clinical chemotherapy drugs and radiation therapy can induce apoptosis. A variety of chemotherapy and radiotherapy agents can directly cause nuclear

28、 DNA damage, which leads to an increase in p53 content, and an increase in p53 levels induces an increase in the apoptotic protein molecule Bax and translocation to the outer mitochondrial membrane. This shift of Bax can interact with Bcl-2 molecules on mitochondria and promote the release of cytoch

29、rome C and apoptosis in mitochondria. P53 can also induce cell cycle dependent activation of the inhibitor P21, causing cell cycle arrest.If DNA damage is repaired, the cell cycle arrest can be relieved, and if the damage cannot be repaired, the cells begin to apoptosis. Mitochondria play a key role

30、 in non receptor mediated apoptosis.3. cell-mediated apoptosis, immune cell killingNot all forms of cell death are cellular suicide. In some cases, a cell can kill another cell. In human and mammalian immune systems, there are T lymphocytes that kill infected cells and other damaged cells. Killer T

31、cells through cell surface receptor recognition of virus infected cells, such as a killer T cells can be worn by Kong Su (Perforin) mediated release of serine protease Granzyme B, cutting a variety of intracellular protein and activation plays an important role in caspases apoptosis, thereby induce

32、cell apoptosis. T lymphocytes also induce apoptosis by releasing large amounts of free radicals via oxygen bursts.Three. The central problem of apoptosis developmentRecently, the study of apoptosis development has focused on the role of mitochondria, the cascade cascade of protease systems (caspase)

33、 and the regulation of various death genes and their products.1. mitochondria: in prokaryotic cells, mitochondria are a small symbiotic bacterium that controls cell life and death in an aerobic environment. The existence of this symbiotic bacterium, from instability to stability and the involvement

34、of nuclear gene spectrum into obligate symbiosis, is a phenomenon of cellular evolution 12. Intracellular oxidative phosphorylation, energy metabolism and anti oxidative activity depends on mitochondrial function. The early changes of mitochondrial mitochondrial membrane permeability changes and mit

35、ochondrial transmembrane potential (mitochondrial transmembrane potential) decreased, open, membrane hole ion by H+ gradient matrix, endometrial damage, osmotic pressure, mitochondrial volume, membrane rupture. The activity of caspases protein released into the cytoplasm by caspases chain reaction;

36、mitochondrial transmembrane potential decreased, uncoupling of respiratory chain, superoxide anion products increase matrix calcium and glutathione efflux, soluble membrane protein, including the Cyto c release, stimulate the cytoplasmic cysteine protease chain reaction, resulting in cell apoptosis

37、or necrosis. Therefore, the size of mitochondrial membrane, the mitochondrial transmembrane potential, and the regulation of apoptosis. There are known 3 mechanisms by which mitochondrial alterations cause cell death: (1) electron transport, oxidative phosphorylation, and destruction by ATP. (2) rel

38、ease proteins that stimulate the caspases family, such as Cyto, C. (3) change the redox potential of cells. The type of death (apoptosis and necrosis) is determined by changes in mitochondria. For example, when stimulating factors such as Bax, Ca2+, and ceramide stimulate mitochondria, there are two

39、 kinds of results, one is osmotic disequilibrium that causes mitochondria to swell, rupture of the outer membrane, electrochemical gradient destruction and ATP depletion, causing cell necrosis. First, increased mitochondrial membrane permeability, release Cyto C, stimulate caspases protein to dissol

40、ve enzymes, produce apoptosis.2.caspases protease: apoptosis is an evolutionary process in which cell suicide is preserved, and the mechanism is protein lysis, system activation, and a family of cysteine proteases known as caspases. The biological characteristics of the caspases family with protease

41、s such as protein dissolution is irreversible; many proteases are from precursor synthesis of the original with little or no catalytic effect, the volume is small, the catalytic role of existence of positive or negative feedback effect; protease can regulate its activity; the corresponding protease

42、inhibitor; the protease reaction is specific. Protease precursor in aspartic acid sites was cut into 3 parts, the H2N- terminal suppression region is removed, the other end of the COOH- end is split into 1 large and 1 small subunits called the death zone, 2 large and 2 small subunits with the surviv

43、al enzyme, can effect caspases downstream of this cascade is called caspases cascade. This chain reaction depends on the regulation of caspases proteins, cofactors, feedback, and thresholds.Aspartic acid specific cysteine protease (caspases) is a homolog of Ced-3 in mammalian cells. This group of pr

44、oteases is structurally related and mainly exists in the cytoplasm of cells. At present, 16 homologous genes have been cloned, one of their common features exist in the form of proenzyme in cells, about 30 50kD, the enzyme originally a large subunit and small subunit domain. In the caspase activatin

45、g enzyme chain, upstream Caspase 8, 9, 10 interact with regulatory proteins such as FADD and / or Apaf to form complexes. Probably the formation of these complexes to caspase auto catalytic cracking, cut off at specific aspartate residues after the peptide bond formation of active molecules. Activat

46、ed upstream caspase can further activate downstream effector molecules (effector, enzymes) such as Caspase, 3, 6, 7, and so on. Not all caspase are involved in apoptosis, such as Caspase 4, 5, 11, 12, 13, and may be involved in inflammatory responses. After caspase is activated, it can cut a wide va

47、riety of intracellular substrates. These specific substrates include: signaling molecules such as MEKK1, p21, activated kinase 2, focal adhesion kinase, Raf-1 and Akt. Apoptosis regulating proteins such as Bcl-2, Apaf-1 and DNA enzymes (DFF, DNA fragmentation factor) inhibitors (ICAD) and so on. The

48、 cleavage of ICAD leads to the release of active DFF and transfer to the nucleus, cutting DNA and forming nucleosomes, which is the main reason for the formation of DNA ladder (DNA, Ladder) which is characteristic of apoptotic cells. The cell cycle regulating proteins, such as Rb (retinoblastoma pro

49、tein, Retinoblastoma, protein), Cyclin, E and so on, can be cut. Structural molecules such as actin, fodrin, gelsolin, lamin, and intracellular important enzymes, such as DNA, repair enzymes, PARP, ADP nucleic acids, sugar, polymerase, Poly (ADP-ribose), polymerase, etc.The Bcl-2 family protein acts

50、 as a checkpoint (check-point) in regulation of apoptosis. Nematode Egl, -1, and Ced-9 genes are cognate analogs of Bcl-2 and its family proteins in humans and other mammalian cells. At present, 24 Bcl-2 family protein genes have been cloned (Figs 8, 3). The family includes anti apoptotic proteins a

51、nd pro apoptotic proteins. Anti apoptotic proteins including Bcl-2, Bcl-w, Bcl-XL and homologous proteins Mcl-1, Al and BHRF1, LMW5-HL, virus encoding protein ORF-16, KS-Bcl-2, ElB-19k, BHl, BH2, and most of them have BH3 and BH4 domains, where BH4 is a domain specific apoptosis inhibitory protein.

52、Pro apoptotic proteins include two subfamilies, namely, the Bax subfamily proteins and the Bid subfamily proteins (containing the BH3 domain). It is thus clear that the BH3 domain is shared by the family and plays a key role in promoting apoptosis. Recent studies have shown that synthetic BH3 short

53、peptides can induce apoptosis in mitochondria.3. gene regulation of apoptosis: there are 3 kinds of apoptosis related genes, genes that promote apoptosis, genes that inhibit apoptosis and genes involved in apoptosis. (1) ced gene 6: CED-3 and CED-4 for nematode apoptosis gene in mammalian animal, si

54、milar to CED-3 gene is ICE, which is a cysteine protease; CED-9 is the inhibitor of apoptosis gene, homologous Bcl-2 and CED-9, can inhibit ced-3. CED-4 is a combining body (adaptor). (2) Bcl-2 and its families: the Bcl-2 family has members of Bcl-x, Bax, Bak, Bad and so on, forming Bcl-2-Bax-Bclx a

55、nd Bax-Bad-Bcl-x regulatory system. Bcl-2 can prevent apoptosis, which is related to tumor cell proliferation and carcinogenesis. Some members, such as Bax, can promote apoptosis (16). (3) ICE gene is a proapoptotic gene in mammals. ICE acts on Pro-IL-1 beta substrates. The gene that inhibits ICE is

56、 Crm A of Cowpox virus, which can induce apoptosis. The Cpp32 gene product may act synergistically with ICE. (4) c-myc and H-ras genes: c-myc promotes apoptosis and H-ras inhibits apoptosis.(5) Fas and Fas ligand (Fas-L): Fas is the death receptor, Fas-L is the product of T lymphocyte, Fas and Fas-L

57、 combine to induce apoptosis. (6) p53 gene: wild type induces apoptosis and inhibits tumor growth through apoptosis. It plays a role in cell cycle and cell damage, which induces apoptosis. (7) Rb has anti apoptotic effect, Rb can arrest cell cycle and interrupt intracellular information transmission

58、. Radiation induced apoptosis may also be mediated by the Rb protein.Four, the possibility of apoptosis in the prevention and treatment of diseaseThe application of caspases theory and explore the mechanism of apoptosis of disease prevention and control method are carried out from two aspects of ani

59、mal experiments and clinical drug research is just unfolding the successful treatment of human cases, but not much, currently used in clinical treatment of the disease include the following categories:Neuronal apoptosis in 1. cerebral ischemia can be prevented by caspases inhibitors. Such as neonatal asphyxia 19, senile cerebral ischemia, Alzheimers disease (such as Alzheimers disease), drugs can be used to prevent neuronal apoptosis.2. preven