Abstract:The task of determining the structure of membrane proteins has been hindered by experimental difficulties. Recent success in crystallising a number of channels (K⁺ channel, Cl^- channel, Aquaporin 1 etc) and pumps (Ca²⁺ pumps) has led to the determination of 3D structures of a number of proteins responsible for selective transport of polar molecules and ions across biological membranes.In recent years several lines of evidence indicate that the lipids in the plasma membrane of animal cells are inhomogeneously distributed. Lipid rafts and caveolae are cholesterol-sphingolipid enriched microdomains. The characteristics of these domains, such as size, composition and dynamics were briefly reviewed in the present paper. A large number of signalling molecules are concentrated within these lipid microdomains, which has been proposed to function as signaling centers. Besides, the roles of lipid rafts in membrane transport were also discussed.

Abstract:Pre-mRNA splicing is a critical step in eukaryotic gene expression and regulation. The removal of introns occurs in spliceosome, a macromolecular machine containing small nuclear ribonucleoproteins (snRNP), heterogeneous nuclear ribonucleoproteins (hnRNP) and K-homologous domain (KH) proteins. Crystallographic and NMR studies have begun to provide insight into the architecture of snRNP and the structural basis for RNA-protein as well as protein-protein interactions involved in pre-mRNA splicing. Elucidating molecular mechanisms underlying pre-mRNA splicing awaits systematic structural analyses of splicing factors and complexes.

Abstract:The subcellular localization and translocation of signaling proteins have risen large interest in the study of cellular signal transduction. MAP kinase pathways are key signaling systems in eukaryotic cells. MAP kinases have relatively specific localization in cells, and translocate into nucleus upon appropriate stimuli, leading to consequent physiological effects. It has been shown that the phosphorylation state of MAPKs as well as the interactions between MAP kinases and other proteins such as the upstream kinases, phosphatases, and downstream substrates may play a role in their specific localization and translocation. The elucidation of the mechanisms of localization and translocation of MAP kinases will be helpful to understand their in vivo functions.

Abstract:Mitochondria, an energy-producing organelle, is involved in the events of growth, development, age, apoptosis and disease. In the development of mammalian embryos produced by nuclear transfer, the change of mitochondria derived from the nucleus donor and the recipient is an interesting and essential question. Based on the research in the laboratory, the recent advances on mitochondria fate in intraspecies and interspecies nuclear transfer embryo were reviewed.

Abstract:Although much progress has been achieved in the last few years, mammalian cloning is still inefficient. Donor cell selection is critical for the success of cloning animal, and it is also an important factor affecting the cloning efficiency. The progress of cloning animal using donor cells of different cell cycle, cell type, cell resource and cell differentiation degree has been summarized.

Abstract:Prostaglandins (PGs) are important in blastocyst implantation and decidualization. The level of PGI₂ was the highest followed by PGE₂ in the site of blastocyst implantation. In addition, PGI₂ level was significantly higher at the implantation sites than inter-implantation sites. IP and PPARs are the G protein-coupled cell surface receptors that are linked to different cytoplasmic signaling pathways and the nuclear hormone receptors, respectively. IP is not expressed or undetectable at implantation sites, but PPARδ expression is very abundant. RXRs (the partner receptor for PPARs), PPARδ-RXRα heterodimer and PGI₂ synthase (COX-2/PGIS) are also abundant at implantation sites. Both PGI₂ agonist (cPGI) and PPARδ specific agonist can restore blastocyst implantation and decidualization in COX-2^－/－ mice. These data suggested that PGI₂ may play important roles in blastocyst implantation and decidualization by PPARδ activation.

Abstract:Centrosome is a tiny organelle which consists of two barrel-shaped centrioles surrounded by a fibrous meshwork termed as the pericentriolar material(PCM). The composition, morphology, size and position of the centrosome in a cell changes continually with cell cycle progression. Duplication of the centrosome is semiconservative and is coordinated with other cell cycle events, including DNA synthesis. Many centrosome-related proteins and kinases have been found to regulate different steps of centrosome duplication. Many growth-suppression genes such as p53, Rb, p21, Gadd45 and Brca1/2 are also involved in control of the duplication process. Centrosome abnormalities are associated with genomic instability and may play important roles in development of human cancers.

Abstract:Transferrin receptor 2 (TfR2) is a newly discovered iron metabolism protein. New findings indicate that in addition to the ability of TfR2 to mediate iron uptake by the liver cells, TfR2 is important in regulation of iron absorption in the small intestine via controlling synthesis and release of hepcidin. The studies have also demonstrated that mutation in the human transferrin receptor 2 gene is one of the causes for hereditory hemochormatosis.

Abstract:The venoms of predatory cone snails represent a rich combinatorial-like library of evolutionarily selected, neuropharmacologically active peptides. A major fraction of the venom components are conotoxins——small, disulfide-rich peptides that potently and specifically interfere with neurotransmission by targeting a variety of proteins expressed on the cell surface, such as the ion channels of Ca²⁺, Na⁺ and K⁺, and the receptors of acetylcholine, 5-hydroxy tryptamine, NMDA, vasopressin and neurotensin. Because of low molecular mass, diversity of structure and targets, high specifiticy and tissue selectivity of conotoxins, they have more advantages over other naturally occurring toxins. Conopeptides could be also used as tools in neuroscience or as therapeutic agents. The conopeptides characterized to date are estimated to represent only 0.1% of the total present in 500 known species of Conus, therefore many novel conotoxins remain to be elucidated and explored. Pharmaceutical companies are now utilizing Conus-derived peptides to develop novel medications for pain, epilepsy and other disorders.

Abstract:It has been shown by researches on mitochondrial genome of higher plant that there are some sequences coming from nuclear or chloroplast genomes, which has been regarded as one basis for higher complexity of mitochondrial genome. Research actuality of horizontal gene transfer among mitochondria, nuclei and chloroplast in higher plant was reviewed.

Abstract:Adipophilin is a specific marker for lipid accumulation in a variety of cells and for diseases associated with fat-accumulating cells. Lipid-laden foam cells derived from macrophages play a critical role in the development of atherosclerosis. By immunohistochemistry with specific monoclonal antibody, it was shown that expression of adipophilin is induced by high-cholesterol-diet feeding in rabbit atherosclerotic lesions. New Zealand white rabbits were fed with high cholesterol chow for 12 weeks. The level of serum total cholesterol, low-density lipoprotein cholesterol, high-density lipoprotein cholesterol, triglyceride, and cholesterol content of aortic wall was investigated. The areas of fatty streak of the aortas was measured after staining with SudanⅣ. The aortic, and liver specimens with HE and immunohistochemistry staining were observed with light microscopes. The level of serum total cholesterol, low density lipoprotein cholesterol, and cholesterol of aortic wall was significantly increased and the areas of fatty streak of the aortas was (40.06±7.29)% at the end of 12-week-cholesterol feeding. The fatty streak of the aorta with immunohistochemistry staining was strongly positive for adipophilin in animals fed with high cholesterol chow, and the liver was negative with or without high cholesterol chow. Antisense oligodeoxynucleotides of mouse adipophilin was also constructed, mouse peritoneum macrophages was cultured with oxLDL or oxLDL plus the antisense fragment. The results showed that adipophilin antisense decreased cellular cholesterol and lipid droplet content of the cell. The data suggested that the expression of adipophilin in vessel walls is related to the hypercholesterolemia, and has a potential role in lipid accumulation in macrophages and pathogenesis of atherosclerosis.

Abstract:Methylenetetrahydrofolate reductase (MTHFR) is one of the important enzymes involved in folate metabolism. A single (C→T) substitution at nucleotide 677 of MTHFR gene influences the enzyme activity and is correlated with susceptibility to several tumour types. In order to compare the association of the MTHFR C677T polymorphism with susceptibility to esophageal squamous cell carcinoma (ESCC) in a Northern Chinese population, high-speed real-time PCR and melting curve analysis were used. The MTHFR C677T genotypes were determined in 189 patients with ESCC and 141 unrelated healthy controls. The results showed that the MTHFR C677T C/C, C/T and T/T genotype frequencies among healthy controls were 17.7%, 38.3%, and 44.0%, respectively. There was no significant difference in MTHFR T/T genotype frequency between ESCC patients (42.3%) and healthy controls (χ²=0.089, P＞0.05) whereas the C/T genotype frequency among ESCC patient (49.2%) was only slightly higher than that among healthy controls (χ²＝3.890, P＜0.05). However, the frequency of the C/C genotype among ESCC patients (8.5%) was significantly lower than that among healthy controls (17.7%) (χ²=6.37, P＜0.05). The C/C genotype significantly reduced the risk for developing ESCC compared to the combination of the C/T and T/T genotypes (OR=0.43, 95% CI=0.22～0.84). The reduced risk was more evident among smokers and patients with family history of upper gastrointestinal cancers. It can be concluded that the MTHFR C677T homozygous wild type may play a protective role in the ESCC development in the Northern Chinese population.

Abstract:In order to investigate whether there exists LMP1/JAK3/ STAT signal pathway in NPC cell line, RT-PCR was first used to detect that four of JAK family members all expressed in two NPC cell lines CNE1 and HNE2. JAK3 which is the most possible associated with LMP1 was used to study further. A stable cell line Tet-on-LMP1 HNE2 expressing LMP1 regulated by tetracycline derivative Dox was used as a model. Western blotting was used to detect JAK3 expression in dose and time dependent fashion under dynamic changes of LMP1. STAT activity was observed after STAT reporter gene GRR-luc was transient transfected in Tet-on-LMP1 HNE2 cell and was induced by varied Dox dose for 36 h. At Dox concentration of 0.06 mg/L, STAT activity reached a peak. JAK3 specific inhibitorⅠ WHI-P131 can inhibit this peak STAT activity at concentration of 3 μmol/L. Therefore, JAK3 expressed in NPC cells can be regulated by LMP1 to activate STAT. The identified LMP1/JAK3/STAT signal pathway maybe plays an important role in NPC carcinogenesis.

Abstract:The three dimensional structure of Arabidopsis thaliana PAP-specific phosphatase was predicted by use of various existing methods on sequence comparison, secondary structure prediction, three dimensional structure prediction and simulation. It was a structure similar with that of Hal2p in Saccharomyces cerevisiae, consisting of an α＋β N-terminal domain and an α/β C-terminal domain. In the predicted structure, possible binding sites for Mg²⁺, as well as for other metal ions, and the structural base sensitive to Na⁺ were found. These sites were related with the biochemical function of Arabidopsis thaliana PAP-specific phosphatase. The structural and functional analysis suggested that the theoretical structure of Arabidopsis thaliana PAP-specific phosphatase, having been deposited in PDB, is not reasonable.

Abstract:To confirm the effect of EBV encoded LMP1 on expression and activation of nuclear transcription factor, Ets-1 and extracellualr signal regulated kinase(ERK) involved in the process. The expression of p-ERK and Ets-1 were assayed with Western blot, and phosphorylation of Ets-1 was assayed with co-immunoprecipitation-Western blot. MEK1/ERK specific inhibitor PD98059 was used to confirm that ERK mediated the activation of Ets-1 by LMP1. The results showed that in nasopharyngeal carcinoma cell line, EBV-LMP1 enhanced the expression of Ets-1 and p-ERK, and phosphorylation of Ets-1 to some extent in time- and dose-dependent manner. With blockade of PD98059, expression of p-ERK induced by LMP1 decreased significantly, and expression and phosphorylation of Ets-1 by LMP1 decreased partly. These results suggest that expression and phosphorylation of Ets-1 by LMP1 was mediated partly by ERK.

Abstract:Activator protein 1 (AP-1) is known to be constitutively activated by the Epstein-Barr latent membrane protein 1 in nasopharyngeal carcinoma cells. Increasing evidence indicated that C-jun N-terminal kinase (JNK), the key upstream kinase of AP-1 mediated signal transduction pathway, plays a key role in the carcinogenesis and progression of nasopharyngeal carcinoma. JNK interacting protein 1 (JIP-1) was newly identified as a potent inhibitor of JNK. The effect of JIP on the proliferation of nasopharyngeal carcinoma cells through interaction with the AP-1 signaling pathway was detected using immunofluroscence, reporter gene, MTT, colony formation and flow cytometric analysis. In nasopharyngeal carcinoma cells, data suggested that JIP down-regulated AP-1 activity through the inhibition of the translocation of phospho-JNK from the cytoplasm to the nucleus. Furthermore, JIP inhibited the rates of cell survival and colony formation. The number of cells in S phase decreased and the number of cells in G1/G0 phase increased after the flow cytometric analysis, suggesting that JIP induced growth arrest of Tet-on-LMP1-HNE2 cells in G1/S phase of the cell cycle. The results, therefore, demonstrated that JIP, by inhibiting AP-1-mediated signal transduction pathway, interfered the cell cycle and may act as an important negative regulator of the proliferation of nasopharyngeal carcinoma cells. Also, it was detected by flow cytometry analysis and laser scanning confocal microscope that JIP triggered the apoptosis of NPC cells. In conclusion, JIP represents a promising new therapeutic molecule for nasopharyngeal carcinoma.

Abstract:Comparative proteomic analysis, combined with 2-DE separation and MALDI-TOF identification, was utilized to compare the protein expression profiles between highly and lowly metastatic subpopulations(i.e.PLA801D and PLA801C) and 11 metastasis-associated proteins were identified and further validated by 1-D Western blotting, Northern blot and/or semi-quantitative RT-PCR analysis. Compared with that in lowly metastatic subpopulation, CK18, TGLC, GDIR, TPMF, IL-18 and ANX1 were significantly up-regulated, while ER60, CH60, PDX1, CLI1 and KCRB were significantly down-regulated in highly metatsatic subpopulation. Most of the candidate proteins have been evidenced to be somehow associated with various aspects of tumor metastasis such as cell growth, motility, invasion, adhesion, apoptosis and tumor immunity, etc. Up to now, CLI1 and IL-18 have not been reported to be associated with lung metastasis, which implied that CLI1 and IL-18 might be new metastasis-associated proteins of lung cancer.

Abstract:In order to develop an alternative and reliable model for further study of human hepatic apolipoprotein CⅢ receptor, and to elucidate the physiological function and regulation of apoCⅢ receptor, whether or not there were apoCⅢ receptors of HepG2 cells by radioligand binding assay was first studied. Addition of increasing concentration of ¹²⁵I-apoCⅢ to HepG2 cells revealed a saturable binding to HepG2 cells with a K_d of （9.53±1.03）×10^-9 mol/L. The maximum specific binding capacity (B_max) was (3.28±0.31) μg/g. The results showed that there were specific, saturable, and reversible binding site of apoCⅢ on HepG2 cells, and it was identified that HepG2 cell line was a useful model for the study of human hepatic apoCⅢ receptor. In the following, the effects of insulin and glucagon on apoCⅢ receptr of HepG2 cells were examined respectively. The results showed that insulin had no effect on the K_d value, but significantly increased the B_max value of apoCⅢ receptor; and glucagon had no effect on the B_max, but decreased the affinity constant of apoCⅢ receptor. These results indicated that human hepatic apoCⅢ receptor was possibliy under the different regulations of insulin and glucagon.

Abstract:To determine the B cell epitope of a monoclonal antibody against the M3-M4 loop of NMDAR1, a random phage displayed dodecapeptide library was screened with the monoclonal antibody MAB363 against the M3-M4 loop of NMDAR1. After four rounds of biopanning，the peptide sequences of positive phage clones were determined and analyzed by DNA sequencing，ELISA and competitive inhibition assay. A positive clone was found (clone1: VHTNPSTWQPIL). The binding between clone1 and MAB363 were competitively inhibited by the recombined M3-M4 loop expressed by E.coli DH5α；The binding between M3M4 and MAB363 could be competitively inhibited by one of solid-synthesized epitope peptides: RLRNPSKD. There were identical sequences among them: NPS. Deleted with NPS, the peptide could not inhibit the binding of MAB363 to M3-M4. These results demonstrate that NPS in M3-M4 loop is the B cell epitope recognized by MAB363，which may be important for developing a practical immunization strategy against excitotoxic brain injury.

Abstract:Cortical spreading depression (CSD) is an important disease model for migraine and cerebral ischemia. The spatio-temporal characteristics of the intrinsic optical signals (IOS) at 540 nm and 660 nm during CSD were investigated by high resolution optical reflectance imaging through thinned skull in rats. The possible physiological mechanisms underlying the IOS were discussed. The CSD were induced by pinprick in 16 α-chloralose/urethane anesthetized Sprague-Dawley rats. A four-phasic IOS response of decreased ［amplitude(－2.1±1.2)%, duration(16.2±3.8)s］, increased ［amplitude(2.9±1.6)%, duration (13.8±2.2)s］, decreased ［amplitude(－14.2±4.5)%, duration(40.6±8.4)s］ and increased (146.2±40.3)s optical reflectance was observed at pial arteries and parenchyma sites in all experimental animals at 540 nm. The IOS spreads peripherally from the site of CSD induction at speeds of (3.7±0.4) mm/min, companying a dramatic dilation (69.2±26.1)% of the pial arteries. At 660 nm, a three-phasic IOS response of increased ［amplitude(3.8±0.6)%, duration(17.9±5.1)s］, decrease ［amplitude(－3.0±1.7)%, duration(43.3±6.4)s］ and increased optical reflectance was recorded at parenchyma sites.

Abstract:The replication-incompetent adenoviral vector containing the cDNA of VEGF₁₂₁ was constructed. After transfection, individual viral plaques were isolated and amplified in HEK293 cells. Confirmed by PCR, both Adeno-X-VEGF₁₂₁ and Adeno-X-LacZ were propagated in HEK293 cells and were purified by CsCl density gradient centrifugation. Adenovirus-mediated VEGF₁₂₁ gene transfer promotes ECV304 proliferation and formation of capillary-like structures in vitro. The expression of VEGF₁₂₁ by adenovirus-infected ECV304 was quantified by enzymelinked immunosorbent assay. Peak VEGF₁₂₁ production was achived at 7～10 days after infection. The conditioned media from ECV304 infected with Adeno-X-VEGF₁₂₁ markedly enhanced vascular permeability. These data may support that adenovirus-mediated VEGF₁₂₁ cDNA gene transfer could provide a useful strategy for efficient delivery of VEGF₁₂₁ in the treatment of ischemic diseases.

Abstract:A novel mouse heart-specific gene, Lrrc10（GenBank Acc No. AF527781）, was cloned from mouse embryo heart by application of EST assembly and RT-PCR. The cDNA of Lrrc10 was 1 410 bp and intronless. This gene was mapped to mouse chromosome 10D2 by BLAST search to mouse genome. The longest ORF of the cDNA encoded a putative proteins of 274 amino acids. Seven leucine rich repeat motifs were present between 53 amino acid to 212 amino acid. No known gene or protein was significantly homologous to Lrrc10 or its deduced protein. However, XM-137268, a predicted human gene submitted by NCBI genome annotation project，shared high identity to Lrrc10. It may be concluded that XM-137268 was the uncloned human orthologue of Lrrc10. BLAST to EST database showed that cDNA of Lrrc10 was supported by 18 ESTs, all of them were from mouse heart. RT-PCR performed on a panel of different mouse tissues demonstrated that expression of Lrrc10 was strongly in heart, low in lung, not or very weakly in other tissues. These results suggest that Lrrc10 is a novel heart specific member of leucine rich repeat superfamily. As far as, no heart-specific member of this family was reported before.

Abstract:O-GlcNAcylation of proteins is a recently discovered post-translational modification of nuclear and cytoplasmic proteins. This modification is similar to protein phosphorylation rather than to classical protein glycosylation of membrane and secreted proteins. Both O-GlcNAcylation and phosphorylation modify the hydroxyl group of serine or threonine residues of tau, the effect of O-GlcNAcylation on phosphorylation of tau was studied. The level of O-GlcNAcylation in differenciated PC12 cells was modulated by changing the concentration of the donor of O-GlcNAcylation and activities of the key enzymes, then, the consequent changes of tau phosphorylation at various phosphorylation sites were examined by using Western blot developed with phosphorylation-dependent and site-specific tau antibodies. It was found that O-GlcNAcylation modulated phosphorylation of tau at many phosphorylation sites and in a site-specific manner. Increased protein O-GlcNAcylation induced a decrease in tau phosphorylation at most of phosphorylation sites, and vise versa. These results suggest that O-GlcNAcylation negatively modulates tau phosphorylation at most phosphorylation sites. Therefore, these studies provide novel insight into the regulation of tau phosphorylation and the molecular mechenism of abnormal hyperphosphorylation of tau in Alzheimer disease brain.

Abstract:The envelope glycoprotein E is a major glycoprotein of pseudorabies virus, which exerts important effect in pseudorabies eradication campaign. The gE gene fragment deleted signal peptide of PRV Fa strain was inserted into Pichia pastoris expression vector pPICZαA, the resulted recombinant expression vector transformed SMD1168 competent cells and obtained engineering Pichia pastoris strain SMD1168/pPICZαA-FL. After high concentration Zeocin^TM screening, phenotype identification, inductive expression, SDS-PAGE and Western blot analysis of culture supernatant, an engineering Pichia pastoris strain SMD1168/pPICZαA-FL-7 in which gE gene fragment deleted signal peptide was expressed in high levels was obtained. SDS-PAGE and Western blot indicated that expression product of gE fragment deleted signal peptide in culture supernatant of SMD1168/pPICZαA-FL-7 was about 80 ku, a little larger than expected. Gel scanning and Bradford protein analysis results showed that expression product reached 11.7 g/L, or 13.49% of total culture supernatant protein in SMD1168/ pPICZαA-FL-7.

Abstract:Chromatin structure plays a critical role in eukaryotic gene expression. Chromatin immunoprecipitation assay (ChIP) provides a powerful tool to analyze the interaction of trans-acting factors with specific chromatin regions in vivo, as well as the role of histone modifications in gene regulation. A simple ChIP protocol is established and used to study the H3 acetylation pattern of the β-globin locus in MEL cells. DMSO induction results in a dramatic increase in H3 acetylation at hypersensitive site HS2 and active gene (βmaj) promoter, whereas the inactive gene (Ey) promoter maintains low acetylation. This result indicates that the ChIP method is feasible.

Abstract:Blue-native polyacrylamide gel electrophoresis is a very powerful procedure that can be used to analyze the components of chloroplast protein complexes. The two photosystems, the ATP synthase, the cytochrome complex b₆f complex, the light-harvesting complexes and the RubisCO are well resolved. Analysis of chloroplast protein on a second gel dimension allows seperation of more than 50 different proteins which are part of multi-subunit enzymes. Western blotting analysis was also used to confirm the identity of the complex. The method has been used to analyze protein complexes of grana and stromal thylakoid.

Abstract:Immobilization of rHir to polyurethane surfaces was studied. A PU1 solution with carboxyl acid in hard segment was cast on PU or Chro surfaces and the carboxyl acid content on the surfaces was measured by methylene blue adhesion. BSA was covalently linked on carboxylated surfaces by EDC and the BSA content on the surfaces was measured by substration of FT-ATR-IR spectra. rHir was immobilized to BSA lay on polymers by EDC and the r-Hir content on surfaces was measured by isotopes tagging. The results showed that the carboxyl acid content on CPU and C-Chro surfaces was 3.77 and 3.03 mmol/m² respectively, the BSA content on CPU-BSA and C-Chro-BSA surfaces was 426 and 262 mg/m² respectively, the rHir content on CPU-BSA-rHir and C-Chro-BSA-rHir surfaces was 716.7 and 691.1 μg/m² respectively. Thrombin activity inhibition test indicated that the KPTT、PT、TT times of rHir-immobilized specimens were prolonged as compared with controlled, the antithrombin activity on rHir-immobilized PU and Chro surfaces was increased obviously and their antithrombogenicity was improved effectively.

Abstract:RNA interference (RNAi) is a biological process in which the introduction of double-stranded RNA (dsRNA) into a cell leads to targeted post-transcriptional gene silencing. Historically, RNAi has been used as a tool for functional genomics research in C.elegans and Drosophila. Initial attempts to activate the RNAi pathway in mammalian cells were unsuccessful, since the introduction of dsRNA＞30 nucleotides in length results in non-specific suppression of gene expression. Much of this response is due to activation of the dsRNA-dependent protein kinase PKR, and the subsequent phosphorylation and inactivation of the translation factor elF2a. As RNAi mechanism being extensively studied, researchers discovered that double stranded short interfering RNA (siRNA) oligotides of 21～23 nucleotides could be used to mediate a gene silencing effect in mammalian cells. The application of RNAi to mammalian cells has the potential to revolutionize the field of functional genomics. The ability to simply, effectively, and specifically down-regulate the expression of genes in mammalian cells holds enormous scientific, commercial, and therapeutic potential.

Abstract:Cytochrome c has been proved to be a powerful scavenger of O2·- and H2O2, the ferricytochrome c scavenges O2·- and the ferrocytochrome c scavenges H2O2. As the respiratory chain leak some electrons from complex Ⅰ and Ⅲ to generate O2·- and then O2·- dismutate to H2O2,the scavenge of O2·- and H2O2 by cytochrome c makes respiratory chain operated with two electron leak bypass. Cytochrome c plays a role to keep O2·- and H2O2 in the normal physiological level in mitochondria. A concept of radical metabolism is introduced into mitochondria based on the electron leak bypass is the metabolic path of O2·-. The role of electron leak of respiratory chain in cell apoptosis is discussed in the dysfunction of radical metabolism of mitochondria.