[Role of glycogen synthase kinase-3ß in development of severe hepatitis liver failure caused by hepatitis B virus].

OBJECTIVE: To investigate the role of glycogen synthase kinase-3ß (GSK3ß) in the development of severe hepatitis liver failure (SHLF) caused by the hepatitis B virus. METHODS: Twelve patients with chronic hepatitis B (CHB) (CHB group), 12 patients with SHLF caused by hepatitis B virus (SHLF group), and 8 normal subjects (control group), who were admitted to Beijing You'an Hospital from January 2009 to December 2011, were included in this study. Their liver tissues were collected to do some clinical examinations. The GSK3ß activity in the liver tissue was detected with a GSK3ß activity assay kit. Western blot was used to determine the expression of p-GSK3, total GSK3, and -actin. The paraffin sections of liver tissue were prepared for immunofluorescence assay. All data were expressed as mean±standard deviation, and comparison between groups was made by least significant difference t test. P < 0.05 was considered statistically significant. RESULTS: Western blot results showed that compared with the control group, the CHB group had a higher level of p-GSK3ß and the SHLF group had a significantly lower level of p-GSK3ß (P = 0.0342). The immunofluorescence assay results showed that the SHLF group had a significantly lower level of p-GSK3ß than the control group. GSK3ß activity assay results showed that compared with the control group, the CHB group had a significantly lower GSK3ß activity and the SHLF group had a significantly higher GSK3ß activity (P = 0.0289), which were consistent with the results of Western blot and immunofluorescence assay. CONCLUSION: GSK3 is activated in the development of SHLF, so it is an important signaling molecule in the pathogenesis of SHLF. Inhibiting its activity may play a role in the prevention and treatment of SHLF.

Outcome of TiNi stent treatments in symptomatic central airway stenoses caused by Aspergillus fumigatus infections after lung transplantation.

BACKGROUND: Central airway stenoses due to Aspergillus fumigatus infections have been a significant cause of morbidity and mortality after lung transplantation. We reviewed our experience using self-expandable braided TiNi-metallic stents in the management of 4 single-lung transplant recipients with central airways stenoses between January 2003 and June 2010. METHODS: Thirty-six single-lung transplant recipients were subjected to pulmonary function testing and surveillance bronchoscopy with biopsy at predetermined intervals and when clinically indicated. Bronchial wash fluid and biopsy material were examined by appropriate fungal stain and culture techniques. RESULTS: Nine of 36 patients (25%) were diagnosed with Aspergillus fumigatus infections; 4 (11.1%) showed rapid decrease in pulmonary function and developed severe upper airway narrowings with about 80% of the central airway obstructed by thick plugs of mucus, heavily laden with Aspergillus species. All 4 patients were managed with stent placement as well as antifungal treatments and showed a forced vital capacity and forced expiratory volume in 1 second improvement of 11.3% and 25.9%, respectively after 1 month. CONCLUSIONS: TiNi stent applications in combination with antifungal drugs are sufficient to treat central airway stenoses after lung transplantation.

Effect of the route of foot-and-mouth disease virus infection of piglets on the course of disease.

Three different routes of Foot-and-mouth disease virus (FMDV) infection of piglets, namely intranasal (i.n.) through drops, intradermal (i.d.) into the foot, and intramuscular (i.m.) were compared regarding the onset and severity of the disease. The results showed that the i.d. injection of the virus resulted in the fastest onset of the disease. The i.m. injection led to a delayed onset, but the final effect was identical with i.d. injection. Moreover, the i.m. injection was simpler to perform and easier to evaluate. Therefore, the i.m. injection of piglets is recommended as the optimal infection route for evaluation of the FMDV vaccine potency.

The complete genome sequence of foot-and-mouth disease virus O/Akesu/58 strain and its some molecular characteristics.

The complete genome of O/Akesu/58 strain of foot-and-mouth disease virus (FMDV) was sequenced. The phylogenetic analysis revealed that it is not closely related to epidemic strains or previous strains compared with reference sequences (the identities of complete VP1 nucleotide sequences range from 77.5 to 84.0%). Its cell-receptor-binding site is a SGD (Ser-Gly-Asp) motif instead of RGD (Arg-Gly-Asp), and 43 bases were deleted in PKs region of the 5'UTR, although deletions were not found in other gene regions.

Molecular cloning and characterization of a factor that binds the human glucocorticoid receptor gene and represses its expression.

The human DNA binding factor GRF-1, which associates with the promoter region of the glucocorticoid receptor gene (hGR gene), is a repressor of glucocorticoid receptor transcription. The GRF-1 cDNA was cloned using polyclonal antibodies against the purified protein. The deduced amino acid sequence from the cDNA sequences show the presence of three sequence motifs characteristic of a zinc finger and one motif suggestive of a leucine zipper in which 1 cysteine is found instead of all leucines. The GRF-1 expressed in COS-1 cells has a molecular weight of 95,000 and enhances the homologous down-regulation of wild-type hGR gene expression. Biochemical analysis suggests that GRF-1 interaction is sequence specific and that transcriptional efficacy of GRF-1 is regulated through its interaction with specific sequence motif, namely 5'-GAAGGAGGTAGCGAGAAAAGAAACTG-GAGAAACTCGGT.GG-3'. The GRF-1 mRNA is 6.5 kilobases long in rat liver and human MCF-7 cells, and its level is regulated by glucocorticoids.

Differential regulation of mouse mammary tumor virus-bacterial chloramphenicol acetyltransferase chimeric gene by human mineralocorticoid hormone-receptor complexes.

The brain tissues of the rat and mouse express two types of corticosteroid binding proteins, the glucocorticoid (GR) and aldosterone (MR) receptors. Unlike the type II (GR) receptor, type I receptor has a high affinity for aldosterone (ALDO) and corticosterone and is structurally similar to the kidney mineralocorticoid receptor (MR). The results reported in this study provide direct evidence for the interaction of dexamethasone (DEX), triamcinolone acetonide (TA), dexamethasone-21-mesylate (DXM) and 11-deoxycorticosterone (DOC) with human MR expressed in cells by transient co-transfection of a hMR expression vector. The interactions of hMR with DEX, TA, DXM, DOC, promegestone (R5020) and methyltrienelone (R1881) were measured by trans-activation of mouse mammary tumor virus long terminal repeat fused to bacterial chloramphenicol acetyltransferase (MMTV-tk-CAT) in gene co-transfection experiments and by cell free hormone binding assay. The incubation of various steroid hormones in the presence of [3H]ALDO in a competition assay with extracts prepared from HeLa cells co-transfected with hMR expression vector, showed that hMR expressed under these conditions has a high relative affinity for DEX which is similar to ALDO, TA and DOC. Incubation with DXM under these conditions showed very little competition, as was observed with R1881 and R5020. Incubation of the co-transfected cells with DEX, ALDO, DOC, R5020, TA, R1881 and DXM demonstrated that the level of trans-activation did not reflect the previously observed order of binding affinity for the hMR. The level of transactivation was always higher with DEX and TA compared to ALDO and DOC. Analysis of the binding of labeled glucocorticoid regulatory element (GRE) and hMR incubated with DEX, ALDO and DXM by gel shift analysis demonstrated that the trans-activation of MMTV-tk-CAT by hMR is a result of the interaction of hMR with GRE in the MMTV-LTR.

Purification of a human glucocorticoid receptor gene promoter-binding protein. Production of polyclonal antibodies against the purified factor.

The glucocorticoid receptor (GR) is an essential protein involved in mediating glucocorticoid-regulated gene transcription. The cellular GR concentration is modulated by a number of factors including glucocorticoids, which are capable of down-regulating their own receptor concentration. To further study this phenomenon, the human GR (hGR) gene promoter was isolated and was shown to contain the sequences essential for glucocorticoid-dependent down-regulation in CV-1 cells by gene transfer. Further transfections performed with the hGR gene demonstrated that the nucleotide sequence between -250 and -750 is implicated in the down-regulation of the hGR by hormone. The promoter region of human GR is extremely rich in G+C sequences which are known to be involved in the regulation of many housekeeping genes such as GR, as well as a number of cellular oncogenes. Using a combination of partial purification of DNA-binding proteins, DNA-protein interaction by gel shift analysis and preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis, we have identified a protein factor (GRF-1) of 95 kDa which interacts with the human GR gene fragment implicated in homologous down-regulation. Polyclonal antibodies were then raised against the protein following purification. This method of purification and identification may be universally applied to purify and characterize unknown factors which may be involved in the regulation of genes such as hGR gene.