Types of Septic Cardiomyopathy: Prognosis and Influencing Factors - A Clinical Study.

Objective: To explore the prognostic outcomes associated with different types of septic cardiomyopathy and analyze the factors that exert an influence on these outcomes. Methods: The data collected within 24 hours of ICU admission included cardiac troponin I (cTnI), N-terminal pro-Brain Natriuretic Peptide (NT-proBNP); SOFA (sequential organ failure assessment) scores, and the proportion of vasopressor use. Based on echocardiographic outcomes, septic cardiomyopathy was categorized into left ventricular (LV) systolic dysfunction, LV diastolic dysfunction, and right ventricular (RV) systolic dysfunction. Differences between the mortality and survival groups, as well as between each cardiomyopathy subgroup and the non-cardiomyopathy group were compared, to explore the influencing factors of cardiomyopathy. Results: A cohort of 184 patients were included in this study, with LV diastolic dysfunction having the highest incidence rate (43.5%). The mortality group had significantly higher SOFA scores, vasopressor use, and cTnI levels compared to the survival group; the survival group had better LV diastolic function than the mortality group (p < 0.05 for all). In contrast to the non-cardiomyopathy group, each subgroup within the cardiomyopathy category exhibited elevated levels of cTnI. The subgroup with left ventricular diastolic dysfunction demonstrated a higher prevalence of advanced age, hypertension, diabetes mellitus, coronary artery disease, and an increased mortality rate; the RV systolic dysfunction subgroup had higher SOFA scores and NT-proBNP levels, and a higher mortality rate (P < 0.05 for all); the LV systolic dysfunction subgroup had a similar mortality rate (P > 0.05). Conclusion: Patients with advanced age, hypertension, diabetes mellitus, or coronary artery disease are more prone to develop LV diastolic dysfunction type of cardiomyopathy; cardiomyopathy subgroups had higher levels of cTnI. The RV systolic dysfunction cardiomyopathy subgroup had higher SOFA scores and NT-proBNP levels. The occurrence of RV systolic dysfunction in patients with sepsis significantly increased the mortality rate.

Early Diastolic Peak Velocity of Mitral Valve Annulus and Right Ventricular Systolic Tricuspid Annular Velocity as Predictors in Assessing Prognosis of Patients with Sepsis.

Objective: To analyze the epidemiological data of patients with septic cardiomyopathy and investigate the relationship between ultrasonic parameters and prognosis of patients with sepsis. Methods: In this study, we enrolled patients with sepsis who were treated at the Department of Critical Care Medicine in the Beijing Electric Power Hospital (No.1 Taipingqiao Xili, Fengtai District, Beijing) from January 2020 to June 2022. All patients received standardized treatment. Their general medical status and 28-day prognosis were recorded. Transthoracic echocardiography was performed within 24 hours after admission. We compared the ultrasound indexes between the mortality group and the survival group at the end of 28 days. We included parameters with significant difference in the logistic regression model to identify the independent risk factors for prognosis and evaluated their predictive value using receiver operating characteristic (ROC) curve. Results: We included 100 patients with sepsis in this study; the mortality rate was 33% and the prevalence rate of septic cardiomyopathy was 49%. The peak e' velocity and right ventricular systolic tricuspid annulus velocity (RV-Sm) of the survival group were significantly higher than those of the mortality group (P < 0.05). Results of logistic regression analysis showed that the peak e' velocity and RV-Sm were independent risk factors for prognosis. The area under curve of the peak e' velocity and the RV-Sm was 0.657 and 0.668, respectively (P < 0.05). Conclusion: The prevalence rate of septic cardiomyopathy in septic patients is high. In this study, we found that the peak e' velocity and right ventricular systolic tricuspid annulus velocity were important predictors of short-term prognosis.

Elevated plasma histone H4 level predicts increased risk of mortality in patients with sepsis.

BACKGROUND: Histones play a vital role in the pathogenesis of sepsis. However, studies on histones and the prognosis of sepsis patients are scarce. This study aims to investigate the relationship between histones and other biomarkers of sepsis. Furthermore, we aim to determine the role histones play in the prognosis of sepsis patients to explore the possibility of using them as a potential biomarker of sepsis. METHODS: We performed a prospective observational study on 136 patients. One hundred twenty-six of them had sepsis, and 10 were enrolled as healthy controls. Baseline blood samples were collected for plasma histone H4, cardiac troponin I (TnI), N-terminal pro-b-type natriuretic peptide (NT-proBNP), procalcitonin (PCT), and lactate. The site of infection, the use of vasopressor, and assessment scores of sequential organ failure were documented within 24 hours of admission. The duration of ICU stay and mortality was also recorded. RESULTS: The mean plasma histone levels of the patients were significantly higher than the healthy controls (P<0.001). Compared with the 89 survivors, the 37 patients who died had a higher rate of sequential organ failure assessment (SOFA) scores (P=0.002), more frequent use of vasopressors (P=0.033), and higher levels of histone H4 (P<0.001). Binary logistic regression analysis showed that high plasma histone H4 levels were independent risk factors for predicting mortality. The area under the receiver operating characteristic curve (0.731) verified that high plasma histone H4 level significantly predicted mortality. Plasma histone H4 levels positively correlated with the SOFA score, and plasma cardiac TnI. CONCLUSIONS: For patients with sepsis in the ICU, an elevated level of plasma histone H4 could be a risk factor associated with an increased mortality rate. Therefore, plasma histone H4 may be a useful biomarker for determining the prognosis of these patients.

Elevated Plasma Histone H4 Levels Are an Important Risk Factor in the Development of Septic Cardiomyopathy

Background: Myocardial impairment is a major complication and an important prognostic predictor of sepsis. Therefore, early and accurate diagnosis as well as timely management of septic cardiomyopathy is critical to achieve favorable outcomes. Aims: To investigate the risk factors of septic cardiomyopathy. Study Design: Cross-sectional study. Methods: This study performed between May 2016 and June 2018 recruited 93 septic patients from the intensive care unit. All patients received standardized treatments. Septic patients were divided into two groups: non cardiomyopathy (n=45) and septic cardiomyopathy group (n=48). Blood samples were collected and transthoracic echocardiography was performed within 24 hours of intensive care unit admission. Septic patients with one ultrasound abnormality but no history of heart disease were diagnosed as having septic cardiomyopathy. Plasma histones, cardiac troponin I, and N-terminal pro-brain natriuretic peptide were measured using ELISA. Sequential Organ Failure Assessment scores, vasopressor use, and the outcomes of intensive care unit stay were analyzed. Spearman rank analysis was used to determine the correlation between plasma histone H4 and other parameters. Binary logistic regression and receiver operating characteristic curve analysis were used to determine the risk factors for septic cardiomyopathy. Results: Compared with the non-cardiomyopathy group, the septic cardiomyopathy group had significantly higher plasma H4 and cardiac troponin I levels, a higher Sequential Organ Failure Assessment score, more frequent vasopressor use, and a higher mortality rate (p<0.05). Plasma histone H4 levels positively correlated with cardiac troponin I (r=0.577, p<0.001), N-terminal pro-brain natriuretic peptide (r=0.349, p=0.001), and Sequential Organ Failure Assessment scores (r=0.469, p<0.001). Binary logistic regression and receiver operating characteristic curve analyses revealed that elevated plasma histone H4 levels and vasopressor use were important risk factors for septic cardiomyopathy (p<0.05). Conclusion: Elevated plasma histone H4 levels could be used to predict septic cardiomyopathy in patients with sepsis.

Improved sepsis bundles in the treatment of septic shock: a prospective clinical study.

BACKGROUND: Sepsis bundles can decrease mortality in patients with severe sepsis or septic shock. However, current methods of measuring pressure, such as central venous pressure, are inadequate. This study investigated the effect of improved sepsis bundles informed by pulse-indicated continuous cardiac output. METHODS: We compared the outcome of treatment with sepsis bundles informed by either conventional pressure measurements or pulse-indicated continuous cardiac output. Patients in 2 groups received fluid resuscitation, standard antibiotics, and oxygen therapy. RESULTS: A total of 105 patients with septic shock were randomly divided into 2 groups: the conventional sepsis bundle group (n = 52) or the improved sepsis bundle group (ISBG, n =53). The ISBG significantly reduced the mean Acute Physiology and Chronic Health Evaluation II and Sepsis-related Organ Failure Assessment scores. Significantly fewer ISBG-treated patients received vasoactive drugs compared to conventional sepsis bundle group-treated patients. In addition, patients in the ISBG exhibited a significantly increased arterial blood lactate clearance rate and required less total fluid resuscitation and a shorter duration of mechanical ventilation and stay in the intensive care unit. CONCLUSIONS: Pulse-indicated continuous cardiac output-directed sepsis bundles can reduce the severity of septic shock, provide more accurate fluid resuscitation, and reduce the duration of mechanical ventilation and stay in the intensive care unit.

[The impact of different fluid management on mortality in patients with septic shock].

OBJECTIVE: To find out the influential effect of different fluid management on mortality of patients with septic shock in different phases. METHODS: From March 2007 to December 2009, a retro spective controlled study was conducted on the clinical data of 107 adult patients with septic shock in the intensive care unit (ICU) of Subei Hospital of Jiangsu Province. The patients were divided into survival group ( n =68) and non survival group ( n =39) according to the final outcome. A number of demographic and variables were collected from the medical record. The acute physiology and chronic health evaluationII (APACHEII) score, sequential organ failure assessment (SOFA), liquid intake and output volume and its balance daily within 1 week, 24 hour early goal directed therapy (EGDT) and conservative late fluid management (CLFM) were compared between two groups. The Logistic regression statistics was used to determine the relationship between APACHEII, SOFA, EGDT, CLFM and survival. RESULTS: The single variable analysis showed that there was significant difference in the parameters of oxygenation index in 7 days ,arterial blood lactate clearance within 24 hours, acute lung injury, length of mechanical ventilation, length of ICU stay and in hospital, the goal of fluid management including 24 hour EGDT, 24 hour CLFM, fluid balance in 24 hours, total fluid input within 7 days, negative fluid balance and times during 7 days between two groups. Logistic regression showed that failure to achieve 24 hour EGDT and late CLFM,a negative balance of <2 000 ml, total fluid input of >20 000 ml within 1 week were independent risk factors of death, and odds ratio ( OR ) was 4.159, 4.431, 23.788 and 4.353, respectively, the P value was 0.035, 0.019, 0.000, 0.025, respectively. The 28 day mortality in 24 hour EGDT and CLFM group (12.5%) was significantly lower than that of 24 hour EGDT with liberal late fluid management (LLFM) group (46.2%) and that in the group of patients in whom with failure to have 24 hour EGDT with CLFM or LLFM (30.0%, 76.2%, P<0.05 or P <0.01). CONCLUSION: Both early achievement of 24 hour EGDT and late CLFM for the patients with septic shock can lower mortality.

[Study of erythromycin and metoclopramide in treatment of feeding intolerance of critically ill patients in intensive care unit].

OBJECTIVE: To compare the effect of erythromycin and metoclopramide on feeding intolerance of critically ill patients in intensive care unit (ICU). METHODS: One hundred and fifty-two critically ill patients in ICU who needed early enteral nutrition exceeding 7 days between January 2007 and January 2009 were included in the study. The patients were randomly divided into three groups: erythromycin group (200 mg intravenous drip, once every 12 hours), metoclopramide group (10 mg intravenous injection, once every 8 hours), and combination therapy group. The whole experiment was carried out for 7 days. Residual gastric volume was aspirated and measured every day at 03:00, 09:00, 15:00 and 21:00. The daily mean gastric residual volume was compared. At the same time, the daily effectiveness of erythromycin and metoclopramide on the success of feeding was also compared. The factors associated with a poor response to prokinetic therapy were looked for. RESULTS: The daily gastric residual volume in the combination therapy group was smallest, the maximum was (40+/-8) ml; the maximum of gastric residual volume in erythromycin group was (42+/-7) ml; the maximum of gastric residual volume in metoclopramide group was (59+/-8) ml (P<0.05 or P<0.01). The successful rate of feeding was highest in the combination therapy group, and it was as high as 97.4%, the erythromycin group ranked the second (90.0%), and that of the metoclopramide group was lowest (89.5%, P<0.05 or P<0.01). Factors that were associated with a poor response to prokinetic therapy was high pretreatment 24-hour gastric residual volume (r=-0.584, P=0.000), high blood sugar level (r=-0.345, P=0.029), a high acute physiology and chronic health evaluation II (APACHEII) score (r=-0.437, P=0.005), and requirement for inotropic drug support (r=-0.389, P=0.041). CONCLUSION: Low dose of erythromycin could improve the successful rate of feeding in critically ill patients in ICU. The combined administration of erythromycin and metoclopramide was more effective. Its side effect was minimal.

[A prospective randomized control clinical study of the effect of Xuebijing injection on prognosis of acute respiratory distress syndrome patients].

OBJECTIVE: To study the effect of Xuebijing injection on prognosis, immune function, adrenal function and inflammatory reaction during the treatment of acute respiratory distress syndrome (ARDS). METHODS: From January 2008 through December 2008, a clinical study was conducted on consecutive adult patients with ARDS in intensive care unit (ICU). The patients were divided into Xuebijing group (31 patients) and control group (30 patients). Both groups were treated with the routine therapy of ARDS, and in addition, Xuebijing injection was used in a dose of 100 ml twice a day for 7 days in Xuebijing group. Duration of mechanical ventilation (MV) and ICU length of stay, 28-day mortality, acute physiology and chronic health evaluation II (APACHE II), Murray and Marshall scores were recorded in both groups. Every patient was given one injection of corticotrophin 250 microg intravenously before and after treatment, and plasma cortisol level was detected by radio-immunoassay before the injection (T0) and 30 minutes (T30) and 60 minutes (T60) after the injection. The ratio of adrenal insufficiency was evaluated according to diagnostic criteria of relative adrenal insufficiency, which was defined as the difference between T0 and the highest value of T30 or T60 (Delta Tmax)< or =248.4 nmol/L. Human leukocyte antigen-DR (HLA-DR), subpopulations of T lymphocyte (CD4(+)/CD8(+)), interleukin-6 (IL-6), IL-10 in peripheral blood was also determined. RESULTS: Murray (1.5+/-1.5) and Marshall score (2.9+/-2.7) and the level of IL-6 [(3.4+/-1.9) micromol/L], IL-10 [(1.5+/-0.8) micromol/L] in the Xuebijing group were decreased significantly after the use of Xuebijing compared with control group [4.3+/-3.1, 6.3+/-4.1, (8.9+/-10.2) micromol/L, (4.2+/-4.8) micromol/L, respectively, all P<0.01], while the values of HLA-DR (41.1+/-10.1), CD4(+) (58.0+/-10.7), CD4(+)/CD8(+) (1.9+/-0.3) were increased compared with control group [30.6+/-15.0, 50.5+/-16.2, 1.4+/-0.7, respectively, P<0.05 or P<0.01]. The ratio of adrenal insufficiency in Xuebijing group (45.2%) was lower than that of control group (83.3%), while that of Delta Tmax [(328.4+/-278.3) micromol/L] was higher than that of control group [(172.8+/-110.8) micromol/L, both P<0.01]. MV duration [(4.0+/-3.3) days] and ICU length of stay [(8.4+/-4.2) days] were less than those of control group [(5.9+/-3.8) days, (12.0+/-7.6) days, both P<0.05], and 28-day mortality in Xuebijing group was 35.5%, which was 11.2% less than that of control group (46.7%), but there was no statistically significant difference between two groups (P>0.05). CONCLUSION: Xuebijing injection improves organ function, decreases MV duration and ICU length of stay in ARDS patients. The underlying mechanism may involve modulation of the immune function, decrease in the degree of adrenal insufficiency, and modulation of regulating inflammatory reaction.

[The impact of sepsis bundles on mortality in patients with sepsis shock: a prospective clinical study].

OBJECTIVE: To survey the impact of sepsis bundle on prognosis of septic shock. METHODS: From January 2007 to June 2008, a prospective clinical study was conducted on consecutive adult patients with sepsis shock in intensive care unit (ICU). The study of sepsis bundle on septic shock was divided into before training (from January to September 2007) and trained phases (from October 2007 to June 2008), and the patients were divided into control group and death group. The relationship between sepsis bundle index and prognosis at 6 hours and 24 hours was analyzed with logistic regression analysis. The independent factors of death due to septic shock were looked for. The compliance of sepsis bundles, duration of mechanical ventilation (MV), ICU stay, and 28-day mortality were noted. RESULTS: One hundred patients, including 51 patients before training and 49 patients after training were enrolled for study. Thirty-six patients survived and 64 patients died. All the patients met the criteria for the diagnosis of sepsis shock. Through analysis of the parameters with logistic regression, it could be found that 6-hour early goal-directed therapy (EGDT) and 24-hour EGDT were the two independent protective factors of death, and standardized regression coefficient was 0.046 and 0.120 respectively (both P < 0.01). Compliance with sepsis bundles was low in training phase, the compliance increased apparently after training. Among them, 6-hour EGDT and 24-hour EGDT compliance increased to 55.1%, 65.3% from 19.6%, 35.3% (both P < 0.01). Compared with the group before training, the duration of MV [(166.6+/-156.4) hours vs. (113.6+/-73.6) hours], that of ICU stay [(9.4+/-7.6) days vs. (6.0+/-3.9) days] and 28-day mortality (72.5% vs. 55.1%, P < 0.05 or P < 0.01) was significantly lower after training. CONCLUSION: Sepsis bundle is able to improve survival rate in patients in septic shock. Compliance with sepsis bundles is low before training, and training can improve the compliance and decrease mortality rate of septic shock.

[A clinical study of subglottic secretion drainage for prevention of ventilation associated pneumonia].

OBJECTIVE: To study the effect of subglottic secretion drainage (SSD) on the incidence of the ventilator associated pneumonia (VAP) in patients receiving mechanical ventilation. METHODS: A retrospective randomly controlled trial was adopted. From January 2005 to June 2006, patients with an expected duration of mechanical ventilation >48 hours and age >18 years were randomly enrolled as SSD group or control group. The following data were analyzed: age, sex, acute physiology and chronic health evaluation II (APACHE II) score, internal diameter (ID) of artificial airway, duration of enteral nutrition (EN), incidence of VAP, duration of mechanical ventilation, length of intensive care unit (ICU) stay, and the mortality in ICU. In SSD group, the amount of secretion was recorded everyday. RESULTS: Sixty-one patients were enrolled in the study, 30 patients were randomized to SSD group, and 31 patients were randomized to control group. The two groups were similar in age, sex, APACHE II score, ID of artificial airway, and duration of EN. Compared with control group, the incidence of VAP was significantly lower (30.0% vs. 51.6%, P<0.05), and the duration of mechanical ventilation [(7.9+/-2.6) days vs. (10.4+/-0.9) days] and length of ICU stay [(9.3+/-2.9) days vs. (12.3+/-5.7) days] were significantly shorter in SSD group (both P<0.05), but the time of onset of VAP [(6.5+/-1.3) days vs. (5.5+/-0.6) days] and mortality (26.7% vs. 38.7%) showed no significant difference between two groups (both P>0.05). In SSD group, if 20 ml was supposed to be regarded as an effective criterion for effective drainage on the first day, the incidence of VAP was 8.7% in 23 patients in whom the criterion was reached, and it was 57.1% in 7 patients in whom the amount of drainage was not up to the criterion (P<0.01). The average daily drainage in patients who developed VAP was significantly lower than those who did not develop VAP [(13.3+/-3.3) ml vs. (37.8+/-11.2) ml, P<0.01]. Among all 61 patients, the duration of mechanical ventilation [(12.1+/-4.5) days vs. (6.9+/-2.5) days], length of ICU stay [(14.0+/-5.3) days vs. (8.4+/-2.5) days], and mortality (52.0% vs. 22.2%) were significantly different between VAP patients and patients without VAP (all P<0.01). CONCLUSION: Effective SSD is one of the important measures to prevent VAP.

[An experimental study on antiviral effects of IFN alpha].

OBJECTIVE: To investigate the effects of different subtypes IFN alpha (IFN alpha2b, IFN alpha2a, and IFN alpha1b) transduction molecular STAT1, STAT2, IFNAR, PKR, and RNase L, and to study the differences of their antiviral effects and to evaluate the key signaling transduction molecules. METHODS: (1) After HepG2 cells were treated with IFN alpha2b, IFN alpha2a, or IFN alpha1b, the mRNA levels of STAT1, STAT2, IFNAR, PKR, and RNase L were detected by RT-PCR. (2) After HepG2 cells were treated with 1000 U/ml IFN alpha2b, IFN alpha2a, or IFN alpha1b, the protein expression levels of STAT1 and IFNAR were examined by Western blot. RESULTS: RT-PCR results: (1) IFNAR, STAT1, and STAT2 mRNA expression levels were slightly higher in the IFN alpha1b group than those in the IFN alpha2b group (P > 0.05). The mRNA expression levels in IFN alpha1b or IFN alpha2b groups were significantly higher than in the IFN alpha2a group (P < 0.05). (2) The PKR mRNA expression showed no significant differences among IFN alpha1b, IFN alpha2b, and IFN alpha2a groups. (3) The RNase L mRNA expression was very weak. We could not compare the differences of the RNase L mRNA levels in different groups by RT-PCR. Western blot results: (1) The IFNAR, and STAT1 protein expressions were greatly up-regulated after IFN alpha induction compared with the untreated group (P < 0.05). (2) The IFNAR, and STAT1 protein expression levels in IFN alpha1b group were slightly higher than the IFN alpha2b group. IFNAR, and STAT1 protein levels of IFN alpha1b or IFN alpha2b group were significantly higher than IFN alpha2a group (P < 0.05). CONCLUSION: STAT1, STAT2, IFNAR mRNA and protein expressions could all be markedly up-regulated after IFN alpha treatment. Effects of IFN alpha1b or IFN alpha2b were greatly stronger than IFN alpha2a. The PKR mRNA expression also was greatly up-regulated after IFN alpha treatment. Expression levels of PKR in IFN alpha1b, IFN alpha2b, and IFN alpha2a groups were all similar. The mRNA level results were consistent with the protein level results. Our results showed that the antiviral activity of IFN alpha1b or IFN alpha2b were stronger than that of IFN alpha2a. The signal transduction molecules STAT1, STAT2, and IFNAR could be regarded as a key index to evaluate antiviral activity of IFN alpha. Further confirmation is still needed to see whether PKR could be regarded as a key index.

Specific anti-viral effects of RNA interference on replication and expression of hepatitis B virus in mice.

BACKGROUND: RNA interference (RNAi) is a powerful tool to silence gene expression post-transcriptionally. Our previous study has demonstrated that small interfering RNAs (siRNAs) have sufficiently inhibited hepatitis B virus (HBV) replication and expression in vitro. In this study we observed the RNAi-mediated inhibitory effects on HBV replication in mice models and accessed the specificity of these effects. METHODS: A mutant RNAi vector (pSI-C mut) with two base pairs different from the original target gene sequence at the RNAi vector (pSI-C) was constructed according to the method described in this study. A mouse model of acute hepatitis B virus infection was established by injecting naked plasmid pHBV1.3 via the tail vein with acute circulatory overload. pSI-C, pSI-C mut and the irrelevant RNAi control plasmid for green fluorescent protein (GFP) gene, pSIGFP were respectively delivered with pHBV1.3 by tail vein injection method. Six days post injection, enzyme-linked immunosorbent assay (ELISA) assay was used to measure the concentration of HBV surface antigen (HBsAg) in mouse serum, immunohistochemical straining method was used to visualize the expression of HBV core protein (HBcAg) in liver tissues, and the transcriptional level of HBV C mRNA in liver tissues was detected by reverse transcriptase PCR (RT-PCR) analysis. RESULTS: Injection of pSI-C exerted magnificent and specific inhibitory effects on the replication and expression of HBV in the murine model. After 6-day post-injection (p.i.), the OD values were shown to be 5.07 +/- 1.07 in infecting group and 0.62 +/- 0.59 in pSI-C group. The concentration of HBsAg in pSI-C group was significantly lower than that in infecting group (P < 0.01). Liver intracellular synthesis of viral core protein was sharply reduced to 0.9% +/- 0.1%, compared with 5.4% +/- 1.2% of positive hepatocytes in infecting group (P < 0.01), and the transcriptional level of HBV C mRNA was greatly reduced by 84.7%. However, the irrelevant RNAi control plasmid (pSIGFP), and the pSI-C mut did not show the same robust inhibitory effects as pSI-C. CONCLUSION: pSI-C exert efficient and specific inhibitory effects on HBV replication and expression in mice models.

[RNA interference inhibits replication and expression of hepatitis B virus in mice].

OBJECTIVE: To develop a mouse model of acute hepatitis B virus infection and to observe the RNA interference-mediated inhibition of HBV replication and expression in the mouse model. METHODS: Thirty Balb/c mice were randomly divided into 5 equal groups: group A to be injected with pHBV1.3, naked plasmid containing 1.3 time HBV ayw type whole genome eukaryotic expression vector, via the caudal vein as infection group; group B to be injected with pHBV1.3 and pSI-C, plasmid expressing HBV-C specific short hairpin RNA, as interference group; group C to be injected with pHBV1.3 and pSI-C mut, a mutant RNAi vector, as mutant interference group; group D to be injected with pHBV1.3 and pGFP, siRNA transcription vector targeting green fluorescence protein (GFP); and group E to be injected with PBS as blank controls. Six days after blood was collected and the mice were killed and their livers were taken out. ELISA was used to measure the concentration of HBsAg in the serum. Immunohistochemistry and RT-PCR were used to detect the expression of HBcAg and HBV C mRNA in the liver. RESULTS: Three days after injection the HBsAg expression in the sera of the infection group was strongly positive. Six days after injection expression of HBsAg was negative in the interference group and blank control group, and was positive in the infection group, mutant interference group, and irrelevant group, however, with significantly lower OD values in the latter 2 groups compared with in the infection group (both P < 0.05). Six days after injection immunohistochemistry showed that HBcAg expression in liver was positive in the infection group, weakly positive in the mutant interference group and irrelevant interference group, and was negative in the blank control group and interference group. RT-PCR showed clear expression of HBV C mRNA in the infection group, mutant interference group, and irrelevant interference. CONCLUSION: RNAi technique specifically and effectively inhibits the replication and expression of HBV. siRNA has significant potential to become a new type antiviral drug. The establishment of an animal model of acute HBV infection in mice by hydrodynamic injection of naked plasmid has solved, to a certain degree, the problem of lack of appropriate animal model of HBV infection.

[Anti-HBV effect of fusion protein (TA1-IFN) in vitro].

OBJECTIVE: To investigate the anti-HBV effect of fusion protein thymosin alpha1-interferon alpha (TA1-IFN) in vitro and to compare its effect with a combination of interferon alpha and thymosin alpha1. METHODS: After 2.2.15 cells were seeded for 24 hours, drugs of five serial concentrations (8000, 4000, 2000, 1000, 500 U/ml) were added to the wells, then the medium was changed every three days. After 2.2.15 cells were treated with drugs for 6 days, the medium was collected. The inhibitory rates on HBsAg and HBeAg were determined using Abbot kit, and the cytotoxicity of different drugs by means of MTT colorimetric assays was also observed. RESULTS: The inhibitory rate of fusion protein on HBsAg, HBeAg was dose-dependent and reached the maximum at 8000 U/ml concentration. In the meantime, the inhibitory rates of fusion protein on HBsAg and HBeAg were 72.2% +/- 0.8% and 60.4% +/- 1.1% respectively, and the cell survival rate was 85.2% +/- 2.0%; In the corresponding concentration, the inhibitory rates of combination thymosin alpha 1 and interferon alpha on HBsAg and HBeAg were 40.0% +/- 0.7%, 34.5% +/- 3.2% respectively. The results showed significant statistical differences between them; cell survival rate 70.0% +/- 1.9%, and the difference of the results was also significant. Cytotoxicity of fusion protein was weaker than a combination of thymosin alpha 1 and interferon alpha. CONCLUSION: Fusion protein TA1-IFN exerted stronger anti-HBV effects in vitro. Its anti-HBV effects in vitro were stronger than the combination of thymosin alpha and interferon alpha, and its cytotoxicity was weaker than the combination of thymosin alpha and interferon alpha. Our studies provided important evidence for clinical research on TA1-IFN, and also brought new hope for hepatitis B therapy.