[Effect of autophagy on liver regeneration].

Autophagy is a lysosome-mediated degradation pathway, which plays an important role in hepatic physiological and pathological processes, in eukaryotic cells. The liver has a remarkable regenerative capacity. After acute or chronic injury, the residual hepatic cells can be activated to enter the cell-cycle for proliferation, in order to compensate for lost liver tissue and recover liver function. In this review, we summarize the relationship between liver regeneration (LR) after various types of injury and autophagy. For example, autophagy is activated to accelerate LR after physically, alcohol and food borne induced liver injury, while the role of autophagy in animal models of LR after chemical injury remains controversial. Autophagy can also be used to promote the replication of virus particles by some hepatotropic viruses (e.g., HBV, HCV) and inhibit LR after viral infection. Studies on mechanisms of autophagy and LR will contribute to clarify the regenerative process and provide new methods for the treatment of liver disease.

Proteomic analysis of the regenerating liver following 2/3 partial hepatectomy in rats.

BACKGROUND: Liver regeneration (LR) after 2/3 partial hepatectomy (PH) is one of the most studied models of cell, organ, and tissue regeneration. Although the transcriptional profile analysis of regenerating liver has been carried out by many reserachers, the dynamic protein expression profile during LR has been rarely reported up to date. Therefore, this study aims to detect the global proteomic profile of the regenerating rat liver following 2/3 hepatectomy, thereby gaining some insights into hepatic regeneration mechanism. RESULTS: Protein samples extracted from the sham-operated and the regenerating rat livers at 6, 12, 24, 72, 120 and 168 h after PH were separated by IEF/SDS-PAGE and then analyzed by MALDI-TOF/TOF mass spectrometry. Compared to sham-operated groups, there were totally 220 differentially expressed proteins (including 156 up-regulated, 62 down-regulated, and 2 up/down-regulated ones) identified in the regenerating rat livers, and most of them have not been previously related to liver regeneration. According to the expression pattern analysis combined with gene functional analysis, it showed that lipid and carbohydrate metabolism were enhanced at the early phase of LR and continue throughout the regeneration process. Ingenuity Pathway Analysis indicated that YWHAE protein (one of members of the 14-3-3 protein family) was located at the center of pathway networks at all the timepoints after 2/3 hepatectomy under our experimental conditions, maybe suggesting a central role of this protein in regulating liver regeneration. Additionally, we also revealed the role of Cdc42 (cell division cycle 42) in the termination of LR. CONCLUSIONS: For the first time, our proteomic analysis suggested an important role of YWHAE and pathway mediated by this protein in liver regeneration, which might be helpful in expanding our understanding of LR amd unraveling the mechanisms of LR.

[Characterization and identification of functional elements in non-coding DNA sequences].

The non-coding DNA sequences refer to non-protein-coding DNA sequences in genome. These sequences can bind with transcription factors or be transcribed as functional RNAs, thus participating in the regulation of many physiological activities and pathological processes. Aiming at gene expression regulation, this review focuses on the recent progress of non-coding DNA and illustrates their structures, functions and potential acting mechanisms. Meanwhile, some computational and experimental methods of identifying functional elements in the non-coding DNAs are introduced. Finally, further studies in this field are proposed.

The role of Kupffer cells in rat liver regeneration revealed by cell-specific microarray analysis.

Liver regeneration after partial hepatectomy is a process with various types of cells involved. The role of Kupffer cells (KCs) in liver regeneration is still controversial. In this study we isolated KCs from regenerating liver and conducted cell-specific microarray analysis. The results demonstrated that the controversial role of KCs in liver regeneration could be explained with the expression patterns of TGF-α, IL-6, TNF, and possibly IL-18 during liver regeneration. IL-18 may play an important role in negative regulation of liver regeneration. The functional profiles of gene expression in KCs also indicated that KC signaling might play a negative role in cell proliferation: signaling genes were down regulated before cell division. Immune response genes in KCs were also down regulated during liver regeneration, demonstrating similar expression profiles to that of hepatocytes. The expression patterns of key genes in these functional categories were consistent with the temporal functional profiles.

The number of the genes in a functional category matters during rat liver regeneration after partial hepatectomy.

Rat liver regeneration after partial hepatectomy (PH) is a good model to study the regulation of cell proliferation. We isolated hepatocytes from regenerating liver at different time points after PH and used microarray Rat Genome 230 2.0 chip to analyze the functional profiles of all up- or down-regulated genes manually and with automatic gene ontological tools. We found that the transcript expressions of PH and sham operation group were apparently different. For PH group, in the priming phase (2-12 h), signaling, transcription, response to stimulus genes predominated in up-regulated genes; in the proliferation phase (24-72 h), cell proliferation genes predominated; in the termination phase (120-168 h), differentiation and translation genes predominated; while metabolism genes predominated in the down-regulated genes at all time points (2-168 h). These functional profiles are consistent with the cellular and molecular phenomenon observed during liver regeneration, and can be closely connected with the biological process. Moreover, the results indicated that not only the quantity of specific genes but also the number of the genes in the specific functional category was regulated during liver regeneration, which means the number of similar genes in a specific functional category matters as well as the regulation of the genes. The changes of the number of the regulated cell proliferation genes and metabolism genes during liver regeneration were similar to the expression patterns of some cell division genes and metabolism genes.

Analysis of time-course gene expression profiles of sinusoidal endothelial cells during liver regeneration in rats.

Liver regeneration (LR) after partial hepatectomy (PH) requires the coordinate contribution of different cell types. Liver sinusoidal endothelial cells (LSECs), representing the largest population of nonparenchymal cells, are proven to be crucial in LR. However, the details about their implications in regeneration are not still clear. In this study, percoll density centrifugation and immunomagentic bead methods were used to isolate LSECs with high purity and yield; global transcriptional profiles of LSECs during the regeneration were investigated by microarray. 1,629 genes were identified to be LR-related. Among them, there were 833 known genes whose expression patterns were clustered into eight classes. Gene function enrichment analysis showed that genes involved in the major LSEC functions, i.e., coagulation, phagocytosis, and transport, were highly enriched in cluster characterized by rapid induction and gradual return, suggesting the quick reestablishment of LSEC function after PH. Genes in immunity/inflammation and defense response were enriched in clusters exhibiting transient downregulation and quick recovery, possibly being associated with suppression of immunity/inflammation pathway in LSECs at early phase. Genes in glycogen synthesis and glycolysis were enriched in the clusters marked by "significant increase and gradual return" and "slight increase and then downregulation", implying an enhanced carbohydrate metabolism at early phase; detoxification-related genes were markedly distributed in the cluster with feature of rapid increase and then reduction, which was helpful in eliminating waste substance. Taken together, the measurement of gene expression profiling of LSECs and expression pattern analysis of functionally categorized genes gave insight into the mechanism of action of this cell on LR.

[Three novel genes BM390716, BI274487 and AA963863 involed in extracellular matrix metabolism of eight rat regenerating liver cell types].

To explore the roles of three novel genes BM390716, BI274487 and AA963863 and the correlation between them during liver regeneration of rats, eight kinds of liver cells were isolated using the combined percoll density gradient centrifugation and immunomagnetic bead method. Rat genome 230 2.0 array was used to detect the changes in expression of genes involved in metabolism of extracellular matrix and the novel genes in rat genome. Correlation between sequence homology, co-expression of the above genes and the physiological activities they involed in were analyzed using Microsoft Excel and BLAST software. The results showed that BM390716 was homologous to and co-expressed with pparα, BI274487 was homologous to and co-expressed with timp2, and AA963863 was homologous to and co-expressed with csgalnact1. It is predicted that BM390716, BI274487, and AA963863 were involved in extracellular matrix metabolism in eight types of rat regenerating liver cells.

[Transcriptome atlas of serine family amino acid metabolism-related genes in eight rat regenerating liver cell types.].

To explore the transcription profiles of serine family amino acid metabolism-related genes in eight liver cell types during rat liver regeneration (LR), eight types of rat regenerating liver cells were isolated using the combination of percoll density gradient centrifugation and immunomagnetic bead methods. Then, the expression profiles of the genes associated with metabolism of serine family amino acid in rat liver regeneration were detected by Rat Genome 230 2.0 Array. The expression patterns of these genes were analyzed through the software of Cluster and Treeview. The activities of serine family amino acid metabolism were analyzed by the methods of bioinformatics and systems biology. The results showed that 27 genes were significantly expressed. Among them, the numbers of genes showing significant expression changes in hepatocytes, biliary epithelial cells, oval cells, hepatic stellate cells, sinusoidal endothelial cells, Kupffer cells, pit cells and dendritic cells were 13, 16, 11, 14, 13, 11, 12, and 14, respectively. The numbers of up-, down-, and up-/down-regulated genes in corresponding cells were 7, 6, and 0; 2, 10, and 4; 2, 8, and 1; 8, 3, and 3; 6, 5, and 2; 4, 6, and 1; 2, 10, and 0; and 6, 6, and 2. Overall, the genes in the eight types of cells were mostly down-regulated during liver regeneration, but most LR-related genes in hepatic stellate cells and sinusoidal endothelial cells were up-regulated in priming phase. It is suggested that biosynthesis of serine family amino acid was enhanced in hepatocytes, hepatic stellate cells, sinusoidal endothelial cells and Kupffer cells in the priming phase. The catabolism of them was enhanced in hepatocytes, biliary epithelial cells, pit cells and dendritic cells in progressive phase.

Possible regulation of genes associated with intracellular signaling cascade in rat liver regeneration.

OBJECTIVE: The importance of signal transduction in cell activities has been generally accepted. The purpose of this study was to analyze the regulatory effect of intracellular signaling cascade-associated genes on rat liver regeneration (LR) at transcriptional level. MATERIAL AND METHODS: The associated genes were originally obtained through a search of the databases and related scientific publications; their expression profiles were then checked in rat LR using the Rat Genome 230 2.0 array. The LR-associated genes were identified by comparing the discrepancy in gene expression changes between the partial hepatectomy (PH) group and the sham operation (SO) group. RESULTS: A total of 566 genes associated with the intracellular signaling cascade were LR related. The genes involved in nine signaling pathways including intracellular receptor-, second messenger-, nitric oxide-, hormone-, carbohydrate-mediated, protein kinase, small GTPase, ER-nuclear and target of rapamycin (TOR) signaling pathways were detected to be enriched in a cluster characterized by up-regulated expression in LR. According to their expression similarity and time relevance, they were separately classified into 5 and 5 groups. CONCLUSIONS: It is presumed that following PH, the second messenger-mediated signaling pathway inhibits the inflammatory response, while the protein kinase cascade and small GTPase-mediated signal transduction stimulate the immune response; the intracellular receptor-, second messenger-, small GTPase-mediated signal transduction and protein kinase cascade coordinately control cell replication; the intracellular receptor-, second messenger-mediated and ER-nuclear signaling pathways facilitate cell differentiation; the MAPK cascade and small GTPase-mediated signal transduction play a role in cytoskeletal reconstruction and cell migration; the second messenger-, small GTPase-mediated and IkappaB kinase/NFkappaB cascades take care of protein transport, etc., in LR.

[Expression profiles of the extracellular matrix-associated genes during rat liver regeneration].

It has been well known that the extracellular matrix (ECM) plays an important role in cell polarization, cell adhesion, cell proliferation, morphogenesis and differentiation. For the sake of the further in-depth investigation of the changes and actions of ECM in liver regeneration (LR) at gene transcriptional level, the ECM-associated genes were obtained by databases searching and literature retrieving, subsequently their expression profiles in rat regenerating liver were detected using Rat Genome 230 2.0 array, then LR-associated genes were identified based on comparison of the gene expression difference between sham operation (SO) group and partial hepatectomy (PH) group. A total of 97 genes were verified to be LR-associated. The initially and totally expressed number of these genes occurring in initial phase of LR, G0/G1 transition, cell proliferation, cell differentiation and structure-functional reconstruction were 49, 19, 43, 5 and 84, 51, 369, 144, respectively, illustrating that expression of the ECM-associated genes were initiated mainly in the early phase, working in differ-ent phases. Their expression similarity was classified into 5 groups including only up-, predominantly up-, only down-, predominantly down-, and equal in up-regulated and down-regulated, involving 38, 21, 21, 10 and 7 genes, respectively; the number of up-regulated expressed genes and down-regulated expressed ones was 411 and 186; their expression patterns were categorized into 24 types, showing that the physiological and biochemical activities in LR were characterized by phase, diversity and intricacy. According to expression profiles and expression patterns of the ECM-associated genes in LR, it was confirmed that the levels of the below-listed genes in expression increased at the corresponding phases of LR, including fibronectin-associated genes at early phase in LR, and collagen-associated genes at middle phase.

Expression patterns and action analysis of genes associated with inflammatory responses during rat liver regeneration.

AIM: To study the relationship between inflammatory response and liver regeneration (LR) at transcriptional level. METHODS: After partial hepatectomy (PH) of rats, the genes associated with inflammatory response were obtained according to the databases, and the gene expression changes during LR were checked by the Rat Genome 230 2.0 array. RESULTS: Two hundred and thirty-nine genes were associated with liver regeneration. The initial and total expressing gene numbers found in initiation phase (0.5-4 h after PH), G(0)/G(0) transition (4-6 h after PH), cell proliferation (6-66 h after PH), cell differentiation and structure-function reconstruction (66-168 h after PH) of liver regeneration were 107, 34, 126, 6 and 107, 92, 233, 145 respectively, showing that the associated genes were mainly triggered at the beginning of liver regeneration, and worked at different phases. According to their expression similarity, these genes were classified into 5 groups: only up-regulated, predominantly up-, only down-, predominantly down-, up- and down-, involving 92, 25, 77, 14 and 31 genes, respectively. The total times of their up- and down-regulated expression were 975 and 494, respectively, demonstrating that the expressions of the majority of genes were increased, and that of a few genes were decreased. Their time relevance was classified into 13 groups, showing that the cellular physiological and biochemical activities were staggered during liver regeneration. According to gene expression patterns, they were classified into 33 types, suggesting that the activities were diverse and complex during liver regeneration. CONCLUSION: Inflammatory response is closely associated with liver regeneration, in which 239 LR-associated genes play an important role.

Correlation analysis of liver tumor-associated genes with liver regeneration.

AIM: To study at transcriptional level the similarities and differences of the physiological and biochemical activities between liver tumor (LT) and regenerating liver cells. METHODS: LT-associated genes and their expression changes in LT were obtained from databases and scientific articles, and their expression profiles in rat liver regeneration (LR) were detected using Rat Genome 230 2.0 array. Subsequently their expression changes in LT and LR were compared and analyzed. RESULTS: One hundred and twenty one LT-associated genes were found to be LR-associated. Thirty four genes were up-regulated, and 14 genes were down-regulated in both LT and regenerating liver; 20 genes up-regulated in LT were down-regulated in regenerating liver; 21 up-regulated genes and 16 down-regulated genes in LT were up-regulated at some time points and down-regulated at others during LR. CONCLUSION: Results suggested that apoptosis activity suppressed in LT was still active in regenerating liver, and there are lots of similarities and differences between the LT and regenerating liver at the aspects of cell growth, proliferation, differentiation, migration and angiogenesis.

Expression patterns and action analysis of genes associated with blood coagulation responses during rat liver regeneration.

AIM: To study the blood coagulation response after partial hepatectomy (PH) at transcriptional level. METHODS: After PH of rats, the associated genes with blood coagulation were obtained through reference to the databases, and the gene expression changes in rat regenerating liver were analyzed by the Rat Genome 230 2.0 array. RESULTS: It was found that 107 genes were associated with liver regeneration. The initially and totally expressing gene numbers occurring in initiation phase of liver regeneration (0.5-4 h after PH), G0/G1 transition (4-6 h after PH), cell proliferation (6-66 h after PH), cell differentiation and structure-function reconstruction (66-168 h after PH) were 44, 11, 58, 7 and 44, 33, 100, 71 respectively, showing that the associated genes were mainly triggered in the forepart and prophase, and worked at different phases. According to their expression similarity, these genes were classified into 5 groups: only up-, predominantly up-, only down-, predominantly down-, up- and down-regulation, involving 44, 8, 36, 13 and 6 genes, respectively, and the total times of their up- and down-regulation expression were 342 and 253, respectively, demonstrating that the number of the up-regulated genes was more than that of the down-regulated genes. Their time relevance was classified into 15 groups, showing that the cellular physiological and biochemical activities were staggered during liver regeneration. According to gene expression patterns, they were classified into 29 types, suggesting that their protein activities were diverse and complex during liver regeneration. CONCLUSION: The blood coagulation response is enhanced mainly in the forepart, prophase and anaphase of liver regeneration, in which the response in the forepart, prophase of liver regeneration can prevent the bleeding caused by partial hepatectomy, whereas that in the anaphase contributes to the structure-function reorganization of regenerating liver. In the process, 107 genes associated with liver regeneration play an important role.

Expression patterns and action analysis of genes associated with drug-induced liver diseases during rat liver regeneration.

AIM: To study the action of the genes associated with drug-induced liver diseases at the gene transcriptional level during liver regeneration (LR) in rats. METHODS: The genes associated with drug-induced liver diseases were obtained by collecting the data from databases and literature, and the gene expression changes in the regenerating liver were checked by the Rat Genome 230 2.0 array. RESULTS: The initial and total expression numbers of genes occurring in phases of 0.5-4 h after partial hepatectomy (PH), 4-6 h after PH (G0/G1 transition), 6-66 h after PH (cell proliferation), 66-168 h after PH (cell differentiation and structure-function reconstruction) were 21, 3, 9, 2 and 21, 9, 19, 18, respectively. It is illustrated that the associated genes were mainly triggered at the initial stage of LR and worked at different phases. According to their expression similarity, these genes were classified into 5 types: only up-regulated (12 genes), predominantly up-regulated (4 genes), only down-regulated (11 genes), predominantly down-regulated (3 genes), and approximately up-/down-regulated (2 genes). The total times of their up- and down-expression were 130 and 79, respectively, demonstrating that expression of most of the genes was increased during LR, while a few decreased. The cell physiological and biochemical activities during LR were staggered according to the time relevance and were diverse and complicated in gene expression patterns. CONCLUSION: Drug metabolic capacity in regenerating liver was enhanced. Thirty-two genes play important roles during liver regeneration in rats.

Expression patterns and action analysis of genes associated with physiological responses during rat liver regeneration: Innate immune response.

AIM: To study the relationship between innate immune response and liver regeneration (LR) at transcriptional level. METHODS: Genes associated with innate immunity response were obtained by collecting the data from databases and retrieving articles. Gene expression changes in rat regenerating liver were detected by rat genome 230 2.0 array. RESULTS: A total of 85 genes were found to be associated with LR. The initially and totally expressed number of genes at the phases of initiation [0.5-4 h after partial hepatectomy (PH)], transition from G0 to G1 (4-6 h after PH), cell proliferation (6-66 h after PH), cell differentiation and structure-function reconstruction (66-168 h after PH) was 36, 9, 47, 4 and 36, 26, 78, 50, respectively, illustrating that the associated genes were mainly triggered at the initial phase of LR and worked at different phases. According to their expression similarity, these genes were classified into 5 types: 41 up-regulated, 4 predominantly up-regulated, 26 down-regulated, 6 predominantly down-regulated, and 8 approximately up/down-regulated genes, respectively. The expression of these genes was up-regulated 350 times and down-regulated 129 times respectively, demonstrating that the expression of most genes was enhanced while the expression of a small number of genes was decreased during LR. Their time relevance was classified into 14 groups, showing that the cellular physiological and biochemical activities during LR were staggered. According to the gene expression patterns, they were classified into 28 types, indicating that the cellular physiological and biochemical activities were diverse and complicated during LR. CONCLUSION: Congenital cellular immunity is enhanced mainly in the forepart, prophase and anaphase of LR while congenital molecular immunity is increased dominantly in the forepart and anaphase of LR. A total of 85 genes associated with LR play an important role in innate immunity.

Expression patterns and action analysis of genes associated with physiological responses during rat liver regeneration: cellular immune response.

AIM: To study the cellular immune response during rat liver regeneration (LR) at a transcriptional level. METHODS: Genes associated with the cellular immune response were obtained by collecting the data from databases and retrieving articles. Gene expression changes during LR were detected by rat genome 230 2.0 array. RESULTS: A total of 127 genes were found to be associated with LR. The number of initially and totally expressing genes in the initial phase of LR [0.5-4 h after partial hepatectomy (PH)], transition from G(0)-G(1) (4-6 h after PH), cell proliferation (6-66 h after PH), cell differentiation and structure-function reconstruction (66-168 h after PH) was 54, 11, 34, 3 and 54, 49, 70, 49 respectively, illustrating that the associated genes were mainly triggered at the initiation of LR, and worked at different phases. According to their expression similarity, these genes were classified into 41 up-regulated, 21 predominantly up-regulated, 41 down-regulated, 14 predominantly down-regulated, 10 similarly up-regulated and down-regulated genes, respectively. The total up- and down-regulated expression times were 419 and 274, respectively, demonstrating that the expression of most genes was increased while the expression of a small number of genes was decreased. Their time relevance was classified into 14 groups, showing that the cellular physiological and biochemical activities were staggered during LR. According to the gene expression patterns, they were classified into 21 types, showing the activities were diverse and complicated during LR. CONCLUSION: Antigen processing and presentation are enhanced mainly in the forepart, prophase and anaphase of LR. T-cell activation and antigen elimination are enhanced mainly in the forepart and prophase of LR. A total of 127 genes associated with LR play an important role in cellular immunity.

Expression patterns and action analysis of genes associated with hepatitis virus infection during rat liver regeneration.

AIM: To study the action of hepatitis virus infection-associated genes at transcription level during liver regeneration (LR). METHODS: Hepatitis virus infection-associated genes were obtained by collecting the data from databases and retrieving the correlated articles, and their expression changes in the regenerating rat liver were detected with the rat genome 230 2.0 array. RESULTS: Eighty-eight genes were found to be associated with liver regeneration. The number of genes initially and totally expressed during initial LR [0.5-4 h after partial hepatectomy (PH)], transition from G0 to G1 (4-6 h after PH), cell proliferation (6-66 h after PH), cell differentiation and reorganization of structure-function (66-168 h after PH) was 37, 8, 48, 3 and 37, 26, 80, 57, respectively, indicating that the genes were mainly triggered at the early stage of LR (0.5-4 h after PH), and worked at different phases. These genes were classified into 5 types according to their expression similarity, namely 37 up-regulated, 9 predominantly up-regulated, 34 down-regulated, 6 predominantly down-regulated and 2 up/down-regulated genes. Their total up- and down-regulation frequencies were 359 and 149 during LR, indicating that the expression of most genes was enhanced, while the expression of a small number of genes was attenuated during LR. According to time relevance, they were classified into 12 groups (0.5 and 1 h, 2 and 4 h, 6 h, 8 and 12 h, 16 and 96 h, 18 and 24 h, 30 and 42 h, 36 and 48 h, 54 and 60 h, 66 and 72 h, 120 and 144 h, 168 h), demonstrating that the cellular physiological and biochemical activities during LR were fluctuated. According to expression changes of the genes, their expression patterns were classified into 23 types, suggesting that the cellular physiological and biochemical activities during LR were diverse and complicated. CONCLUSION: The anti-virus infection capacity of regenerating liver can be enhanced and 88 genes play an important role in LR.

Identification of expressed genes in regenerating rat liver in 0-4-8-12 h short interval successive partial hepatectomy.

AIM: To identify the genes differentially expressed in the regenerating rat liver of 0-4-8-12 h short interval successive partial hepatectomy (SISPH) and to analyze their expression profiles. METHODS: Five hundred and fifty-one elements screened from subtractive cDNA libraries were made into a cDNA microarray (cDNA chip). Extensive gene expression analysis following 0-4-8-12 h SISPH was conducted by microarray. RESULTS: One hundred and eighty-three elements were selected, which were either up- or down-regulated more than 2-fold at one or more time points after SISPH. Cluster analysis and generalization analysis showed that there were five distinct temporal patterns of gene expression. Eighty-six genes were unreported, associated with liver regeneration (LR). CONCLUSION: Microarray analysis shows that the down regulated genes are much more than the up-regulated ones in SISPH; the numbers of genes expressed consistently are fewer than that expressed immediately; the genes expressed in high abundance are much fewer than that increased 2-5-fold. The comparison of SISPH with partial hepatectomy (PH) shows that the expression trends of most genes in SISPH and in PH are similar, but the expression of 43 genes is specifically altered in SISPH.

Expressed genes in regenerating rat liver after partial hepatectomy.

AIM: To reveal the liver regeneration (LR) and its control as well as the occurrence of liver disease and to study the gene expression profiles of 551 genes after partial hepatectomy (PH) in regenerating rat livers. METHODS: Five hundred and fifty-one expressed sequence tags screened by suppression subtractive hybridization were made into an in-house cDNA microarray, and the expressive genes and their expressive profiles in regenerating rat livers were analyzed by microarray and bioinformatics. RESULTS: Three hundred of the analyzed 551 genes were up- or downregulated more than twofolds at one or more time points during LR. Most of the genes were up- or downregulated 2-5 folds, but the highest reached 90 folds of the control. One hundred and thirty-nine of them showed upregulation, 135 displayed downregulation, and up or down expression of 26 genes revealed a dependence on regenerating livers. The genes expressed in 24-h regenerating livers were much more than those in the others. Cluster analysis and generalization analysis showed that there were at least six distinct temporal patterns of gene expression in the regenerating livers, that is, genes were expressed in the immediate early phase, early phase, intermediate phase, early-late phase, late phase, terminal phase. CONCLUSION: In LR, the number of down-regulated genes was almost similar to that of the upregulated genes; the successively altered genes were more than the rapidly transient genes. The temporal patterns of gene expression were similar 2 and 4 h, 12 and 16 h, 48 and 96 h, 72 and 144 h after PH. Microarray combined with suppressive subtractive hybridization can effectively identify the genes related to LR.

iTRAQ-based proteomic analysis of adaptive response in the regenerating limb of the Cynops orientalis newt.

The newt has the powerful capacity to regenerate lost limbs following amputation, and represents an excellent model organism to study regenerative processes. However, the molecular basis of the adaptive response in the regenerating limb of the Chinese fire-bellied newt Cynops orientalis immediately after amputation remains unclear. To better understand the adaptive response immediately after limb amputation at the protein level, we used isobaric tags for relative and absolute quantitation (iTRAQ) coupled with LC-MS/MS methods to analyze changes in the proteome of the regenerating newt limb that occurred 2 h and 8 h after amputation. We identified 152 proteins with more than 1.5-fold change in expression compared to control. GO annotation analysis classified these proteins into several categories such as signaling, Ca(2+) binding and translocation, transcription and translation, immune response, cell death, cytoskeleton, metabolism, etc. Further ingenuity pathway analysis (IPA) showed that several signaling pathways were significantly changed at 2 h and 8 h after amputation, including EIF2 signaling, acute phase response signaling, tight junction signaling and calcium signaling, suggesting these pathways may be closely related to the adaptive response immediately after limb amputation. This work provides novel insights into understanding the molecular processes related to newt limb regeneration immediately after amputation, and a basis for further study of regenerative medicine.