Diosgenin Glucoside Protects against Spinal Cord Injury by Regulating Autophagy and Alleviating Apoptosis.

Spinal cord injury (SCI) is a severe traumatic lesion of central nervous system (CNS) with only a limited number of restorative therapeutic options. Diosgenin glucoside (DG), a major bioactive ingredient of Trillium tschonoskii Max., possesses neuroprotective effects through its antioxidant and anti-apoptotic functions. In this study, we investigated the therapeutic benefit and underlying mechanisms of DG treatment in SCI. We found that in Sprague-Dawley rats with traumatic SCI, the expressions of autophagy marker Light Chain 3 (LC3) and Beclin1 were decreased with concomitant accumulation of autophagy substrate protein p62 and ubiquitinated proteins, indicating an impaired autophagic activity. DG treatment, however, significantly attenuated p62 expression and upregulated the Rheb/mTOR signaling pathway (evidenced as Ras homolog enriched in brain) due to the downregulation of miR-155-3p. We also observed significantly less tissue injury and edema in the DG-treated group, leading to appreciable functional recovery compared to that of the control group. Overall, the observed neuroprotection afforded by DG treatment warrants further investigation on its therapeutic potential in SCI.

[Research progress of mitochondrial fission and fusion related proteins].

Mitochondria are an essential component of multicellular life and play important roles in the health of the cells and the body. Mitochondria can produce energy by oxidative phosphorylation, mediate calcium and reactive oxygen signal transduction, and regulate cell apoptosis. Recent studies indicate that mitochondria continually change their shapes and distribution by fission and fusion, which are collectively termed mitochondrial dynamics. Mitochondrial dynamics play critical roles in maintaining mitochondrial function. This review focuses on the structure and biological functions of mitochondrial fission and fusion related proteins in mammal cells.

Biological characteristics of the rtA181T/sW172* mutant strain of Hepatitis B virus in animal model.

BACKGROUND: The effects of Hepatitis B virus (HBV) rtA181T/sW172* mutation on viral replication and pathogenicity was concerned recently. This study aimed to investigate the biological characteristics of rtA181T/sW172* mutant strain of HBV in animal model. METHODS: The rtA181T/sW172* mutant plasmid was constructed using the pHBV4.1 (wild type HBV) as a template. The wild and mutant HBV replication mouse models were established utilizing a hydrodynamic technique. The titers of hepatitis B surface antigen (HBsAg), hepatitis B e antigen, and HBV DNA in serum, and the levels of HBsAg, hepatitis B core antigen(HBcAg), HBV DNA replication intermediates (HBV DNA RI) and HBV RNA in liver were measured after 1, 3, 5, 7, 10, 12 and 15 days of plasmid injection. RESULTS: In wild-type HBV replication mouse model, serum HBsAg was high on day 1, 3, and 5, but became lower since day 7; while in mutant HBV mouse model, serum HBsAg was always at very low level. In liver tissues, HBV DNA RI of wild type HBV was detected on day 1 after transfection. The level subsequently peaked on day 3, gradually declined after day 5, and was almost undetectable on day 10. However, the HBV DNA RI levels of the mutant strain were always higher and lasted longer until day 15. Consistently, the expression levels of HBsAg and HBcAg in liver of the mutant group were significantly increased. CONCLUSIONS: In the case of the HBV rtA181T/sW172* mutation, the secretion of serum HBsAg was impaired, whereas HBV DNA replication and HBsAg/HBcAg expression were increased in liver. These results suggest that the mutation can impair HBsAg secretion, and may cause the accumulation of viral core particles in liver.

Study of the expression levels of Hepatocyte nuclear factor 4 alpha and 3 beta in patients with different outcome of HBV infection.

Hepatocyte nuclear factors 4 alpha (HNF4α) and 3 beta (HNF3ß) are members of a group of liver-enriched transcription factors (LETFs) that play important roles in regulating the replication of hepatitis B virus (HBV) and liver inflammation. However, the relationship of the level of HNF4α and HNF3ß with the severity of HBV-infected liver diseases is unclear. In this study, liver tissue samples from different types of HBV patients were collected, and HNF4α and HNF3ß expression were detected by immunohistochemistry. The expression of HNF4α was significant higher in patients with severe hepatitis B(SHB) than those with chronic hepatitis B(CHB) and liver cirrhosis(LC) (both P < 0.05), but similar between patients with CHB and LC (P > 0.05). And the expression of HNF3ß was similar among patients with CHB, LC and SHB (P > 0.05 for all pairwise comparison). This suggests that the expression level of HNF4α was different in patients with different outcome of HBV infection, high expression level of HNF4α may correlate with occurrence of SHB.

Histological changes in chinese chronic hepatitis B patients with ALT lower than two times upper limits of normal.

The role of ALT as a predictor of liver injury has been questioned. The aim of this study is to use liver biopsy to assess the degree of liver injury in patients with chronic hepatitis B(CHB) whose ALT < 2 x upper limit of normal (ULN). A total of 49.2% of patients in this study had significant inflammation (grade >or=2) and 36.4% had significant fibrosis (stage >or=2). The frequency of serious inflammation and fibrosis was similar in patients with different ALT levels. The level of serum HBV DNA was not significantly associated with the extent of inflammation and fibrosis. Advanced age was a significant independent predictor of histological damage and the presence of more significant inflammation and fibrosis. We conclude that many CHB patients with ALT < 2 x ULN have significant liver inflammation or fibrosis and that liver biopsy is necessary to assess liver damage and should be used to assess the need for anti-viral therapy.

[Forensic pathological analysis on 23 cases of fatal pulmonary thromboembolism].

OBJECTIVE: To analyze the forensic pathological characteristics of sudden death caused by pulmonary thromboembolism and the chronological transformation of thrombus and explore the assessment method of the causal relationship between previous trauma and the following fatal PTE episode. METHODS: All the 23 cases reviewed here were collected from our institute files from the year of 1998 to 2008. RESULTS: Trauma, surgery and braking etc. were all risky factors of PTE. Of these cases, 12 cases were caused by trauma, 21 cases were caused by surgery and 22 cases died in hospitals which were often happened one or two weeks after injury or one week's postoperative time. Of all the cases, 6 cases had single attack of thrombus and the rest 17 cases had the recurrence of thrombus. The number of the leg deep vein to be the embolic source was 16 cases which were often seen in the left leg. CONCLUSION: It is important to confirm the embolic source, trauma, surgery and chronological events in determing the sudden death with PTE.

Establishment of cell lines using a doxycycline-inducible gene expression system to regulate expression of hepatitis B virus X protein.

The hepatitis B virus (HBV) X gene plays an important role in HBV-associated pathogenesis, especially hepatocarcinogenesis. Establishment of a stable and regulable HBx expression system will allow study of the function of this gene. Here, we describe the development of a doxycycline-inducible recombinant plasmid (pBPSTR3-FlagX) with the full-length HBV X gene and all components of the tetracycline-on ("Tet-on") gene expression system. This vector exhibited dose-dependent doxycycline-dependent induction of the Flag-HBx protein in HepG2 and Hep3B cells. We also observed dose-dependent doxycycline transactivation of HBx in HepG2 cells. After transfecting HepG2 cells with the pBPSTR3-FlagX plasmid, we isolated five puromycin-resistant cell clones with stable HBx expression, two of which exhibited stable and tight control of HBx expression by doxycycline. This new system has great potential for functional studies of the HBV X gene.

[Analysis of death caused by postpartum hemorrhage].

OBJECTIVE: To analyze the pathological characteristics and the death reasons due to postpartum hemorrhage, and to help to deal with the obstetrical medical tangles. METHODS: Thirty-two cases of death caused by postpartum hemorrhage encountered in our department since 1995 had been collected and retrospectively analyzed. RESULTS: Death caused by postpartum hemorrhage could be divided into single factor and multi-factor, with 81.25% due to single factor, 12.50% multi-factor, and 6.25% unknown reason. The single factors included uterine atony, retained placenta, placenta increta, laceration of the lower genital tract, and coagulation defects. The multi-factor included a combination of two or more factors mentioned above. CONCLUSION: The causes of death due to postpartum hemorrhage should be analyzed according to the clinical characteristics of the postpartum hemorrhage and the autopsy examination.

[Expression of myocardial collagen I in early acute myocardial ischemic of human].

OBJECTIVE: To estimate the significance of myocardial collagen I in the early acute myocardial ischemia of human. METHODS: The myocardial paraffin block had normal group, early myocardial ischemia group and myocardial infarction group. The myocardial collagen I was observed with immunohistochemical staining and analyzed by half-quantity. RESULTS: The similar expression of collagen I was observed in the cytoplasm and nucleolus in two groups. CONCLUSION: The collagen I appeared in the myocardium of early acute myocardial ischemia of human. It may be an important index for postmortem diagnosis of the early acute myocardial ischemia.

[Effects of ginsenoside Rg1 on expression of insulin-like growth factor-1 in brain of rats with brain contusion].

OBJECTIVE: To study the effects of ginsenoside Rg1 on the expression of insulin-like growth factor-1 (IGF-1) in the brain of rats after the experimental brain contusion. METHODS: A total of twenty-six Wistar rats were randomly divided into normal control group (n=2), untreated group (n=8) and ginsenoside Rg1 group (n=16). Brain injuries were induced in rats by a mechanical striking device. The brain tissues were extracted at different times after brain injury (6th hour, 12th hour, 2nd day, 6th day), then the expression of IGF-1 in brain tissue was examined by immunohistochemical method. RESULTS: In comparison with the normal control group, the expression of IGF-1 in the brain tissues was increased in the untreated group after the brain contusion (P<0.05). The expression of IGF-1 in brain tissues in ginsenoside Rg1 group was significantly increased as compared with the untreated group (P<0.05). CONCLUSION: Ginsenoside Rg1 enhances the recovery of the contused brain through increasing the expression of IGF-1.

[Sex determination by studying head CT film].

OBJECTIVE: To explore the methods for sex determination on head CT film. METHODS: To establish the sex determination equations by binary regression analysis of cranial skeletal indices obtained on head CT film. RESULTS: A single element equation by applying the index of the width of ansa capitis and a multi-element equation by applying multitude indices were established. CONCLUSION: The equations for sex determination on head CT film were established.

[On the role of liver-enriched transcription factors in regulating HBV transcription and replication].

OBJECTIVE: To investigate the effects of various liver-enriched transcription factors in regulating HBV transcription and replication, and to explore their potential roles in HBV hepatotropism. METHODS: The replication-competent HBV recombinant plasmid pHBV4.1 plus different liver-enriched transcription factor (HNF1, HNF3, HNF4, HNF6, C/EBP and RXRa/PPARa) expression plasmids were co-transfected into nonhepatic cell lines (NIH3T3, HeLa, 293T, SW1353, CV-1 and COS1). The transcription levels of 3.5 kb, 2.4/2.1 kb and 0.7 kb HBV RNA were analyzed by Northern blot hybridization, and the level of HBV DNA replication intermediates was detected by Southern blot hybridization analysis. RESULTS: In the absence of co-transfected liver enriched transcription factor expression vectors, the 3.5 kb HBV RNA is not transcribed and HBV DNA replication is not detected after transfecting of NIH 3T3 cells with pHBV4.1. Expression of the liver-enriched transcription factor HNF4 or RXRalpha/PPARalpha, stimulates the transcription of 3.5 kb HBV RNA and the replication of HBV DNA. In contrast, expression of HNF1, HNF3, HNF6 and C/EBP does not stimulate the transcription of 3.5 kb HBV RNA and therefore does not activate viral replication. HNF4 and RXRalpha/PPARalpha were also shown to activate the transcription of 3.5 kb HBV RNA and viral replication in divers cell types including HeLa, 293T, SW1353, CV-1 and COS1 cells. Mutation of the proximal nucleocapsid HNF4 binding site results in a greatly decreased level of HNF4 or RXRalpha/PPARalpha dependent HBV replication. CONCLUSION: This study demonstrated that the liver-enriched transcription factors HNF4 and RXRa/PPARa can support HBV transcription and replication in nonhepatic cells, indicating that liver-specific gene transcription is one of the determinants of HBV hepatotropism.

[Comparative study for the methods on measuring intracranial hemorrhage volume].

OBJECTIVE: To explore a method of measuring intracranial hemorrhage to be applied in clinical forensic science. METHODS: The accurace of the methods of Tada formula, Stereology and Software boundary to measure intracranial hemorrhage and their practicability were compared. RESULTS: The measured results of Stereology and Software boundary are precise, but the irregularity haematoma volume measured by the method of Tada formula has a high error value. CONCLUSION: The method of stereolgy is the best in measuring intracranial hemorrhage for clinical forensic science.

Biological characteristics of the A1762T/G1764A mutant strain of hepatitis B virus in vivo.

The double nucleotide, A1762T and G1764A exchange (TA mutation), in the hepatitis B virus (HBV) genome basal core promoter (BCP) region is a common viral mutation in patients with chronic HBV infection. This mutation is located in the binding site of hepatocyte nuclear factor 4 (HNF4), and a number of liver­enriched transcription factors are involved in the regulation of HBV transcription and replication. The aim of the present study was to investigate the biological characteristics of the HBV strain with this mutation, and the effect of HNF4 inhibition on the replication of this strain in vivo. The results indicated that in vivo the HBV strain with the TA mutation supported a higher level of pregenomic RNA transcription and HBV DNA replication, compared with the wild­type strain. Furthermore, the concentration of serum HBeAg in the TA mutant group was lower than that in the wild­type strain. Following treatment of the mice with entecavir (ETV) or tenofovir disoproxil fumarate (TDF), the transcription and replication levels of wild­type and mutant strains were reduced. In the groups treated with TDF, the inhibition effect was more marked. In hepatocytes in which HNF4 expression was specifically inhibited, the level of 3.5 kb mRNA of HBV was reduced compared with that in mouse cells with normal HNF4 expression, and HBV DNA replication levels were also reduced to a greater extent. Furthermore, following liver­specific knockdown of HNF4, the reduction in variant virus expression was greater than that of the wild­type virus. In conclusion, the replication capacity of HBV with the TA mutation was increased, and the mutation was associated with a reduction in serum HBeAg levels. This mutant strain remained sensitive to ETV and TDF, and HNF4 supported a higher replication level of TA mutant HBV in vivo.

Role and functional domain of hepatitis B virus X protein in regulating HBV transcription and replication in vitro and in vivo.

The role of hepatitis B virus (HBV) X protein (HBx) in the regulation of HBV replication remains controversial. In the present study, the role of HBx in regulating HBV replication was initially investigated in both HepG2 and Huh7 in vitro cell lines with a transient transfection system. Next, the regions of HBx responsible for transcriptional transactivation and promotion of HBV replication were mapped in an HBV replication mouse model by in vivo transfection of a series of HBx expression plasmids. In an in vitro setting, HBx deficiency had little effect on HBV replication in Huh7 cells, but impaired HBV replication in HepG2 cells. In an in vivo setting, HBx had a strong enhancing effect on HBV transcription and replication. For the C-terminal two-thirds of the protein (amino acids [aa] 51 to 154) was required for this function of HBx, and the regions spanning aa 52 to 72 and 88 to 154 were found to be important for the stimulatory function of HBx on HBV replication. In conclusion, the role of HBx in HBV replication regulation is affected by host cell type, and HBx has an important role in stimulating HBV transcription and replication in hepatocytes in vivo. Further, the transcriptional transactivation function of HBx may be crucial for its stimulatory effect on HBV transcription and replication.