OBJECTIVETo investigate the effect of hepatitis B virus (HBV) X protein (HBx) on host cell cycle and HBV replication using cultured primary mouse hepatocytes to gain further insights into the mechanism of HBx-mediated modulation of cell cycle.

METHODSPrimary cultured mouse hepatocytes were transfected with HBx-expressing (pHBV) or HBx-selected (pHBV triangle X) plasmids, which were generated with sequences of the HBV ayw subtype 1.2 and included the green fluorescent protein (GFP) reporter gene. The levels of cell cycle proteins (p16, cyclin D1, p21, cyclin E and cyclin A) were measured with Western blotting, and HBV DNA was analyzed by Southern blotting and real-time PCR.

RESULTSThe freshly isolated hepatocytes showed no significant differences in levels of cell cycle proteins. However, at 48 hours post-transfection, the levels of cyclin D1, p21 and cyclin E were significantly higher and the level of p16 was significantly lower in the pHBV-transfected hepatocytes than in the pHBV triangle X-transfected hepatocytes (t = 15.713, 22.897, 14.680, and -19.584, respectively, P less than 0.05). The level of cyclin A was not different between the two groups (t = 0.142, P more than 0.05). At 72 hours post-transfection, the level of HBV DNA was higher in pHBV-transfected hepatocytes (rcDNA: 3288.336+/-448.011; dslDNA: 6458.318+/-182.163; ssDNA: 2760.613+/-393.561) than in pHBV triangle X-transfected hepatocytes (rcDNA: 515.721+/-62.530; dslDNA: 2122.228+/-28.347; ssDNA: 1632.013+/-207.021) and in the blank (untransfected) control group (P less than 0.05). Real-time PCR analysis of HBV DNA copy number per cell confirmed these results, (pHBV-transfected: 987.50+/-47.80 vs. pHBV triangle X-transfected: 303.67+/-33.94; t = 20.203, P less than 0.05).

CONCLUSIONSThe HBx protein can affect the levels of cell cycle proteins, which may induce quiescent hepatocytes to enter the G1 phase of the cell cycle and stay in this phase instead of entering the S phase, thereby promoting HBV intracellular replication.

Animals ; Cell Cycle ; Cell Cycle Proteins ; genetics ; metabolism ; Cell Line ; Hepatocytes ; cytology ; virology ; Male ; Mice ; Mice, Inbred C57BL ; Plasmids ; Trans-Activators ; genetics ; metabolism ; Transfection

Objective To study the value of carotid plaque magnetic resonance imaging (MRI) in pre-operation assessment in the elderly patients with carotid atherosclerosis and explore the possibility of minimizing the contrast weightings to gain sweeptime. Methods Totally 70 elderly patients with cerebral ischemia (average age of 68.8 years) underwent carotid MRI and digital subtraction angiography (DSA) due to the appearance of carotid plaque detected by ultrasound. Carotid plaque MRI was acquired with 3.0T MR scanner and 8 channel surface coil. The standard carotid plague MRI program included pre-and post-contrast T1 weighted imaging (T1WI), T2 weighted imaging, proton density weighted imaging and 3D time of flight MR angiography (3D TOF MRA). All these program were divided into two combinations: the 5-sequence MRI (all the sequences) and 2-sequence MRI (T1WI and TOF MRA). Digital subtraction angiography (DSA) in coronal and lateral views of carotid artery was performed with GE Advantx LCN+. The software SPSS 13.0 was used to statistically analyze the difference between MRI and DSA, and that of two sequence combinations was used in the detection of luminal stenosis and fibrous cap (FC) rupture. Results Totally 135 arteries were analyzed while 3 arteries in one patient were excluded due to the poor quality image and stent placement. The degree of luminal stenosis were (38.3±31.0)% and (38.5±30.9)%, respectively, detected by the two MRI sequence-combination with no significant difference (t=2.447, P＞0.05) and was (35.1±31.8)% by DSA. There was a good concordance between MRI and DSA in luminal stenosis detection (Kappa value: 0.773). No statistical difference was found between two MR sequence combinations in detecting FC rupture (both in 36 vessels). DSA detected FC rupture of 16 vessels, showing remarkably difference contrast to MRI(χ2=12.0, P＜0.01). Conclusions MRI can accurately detect the luminal stenosis and FC rupture. The short time scanning resulting from sequence optimization could make MRI much more suitable than DSA to do the pre-operation assessment for senile carotid atherosclerotic patients.

Objective To investigate the significance and safety of repeat transurethral resection(Re‐TUR) for treating stage T1 of non‐muscle invasive bladder cancer .Methods The clinical data were retrospectively analyzed on 41 cases of stage T1 of non‐muscle invasive bladder cancer in this department of our hospital from January 2013 to November 2014 .All cases underwent Re‐TUR at 4-6 weeks after primary surgery .Among them ,33 cases were male and 8 cases were female ,24 cases were single tumor and 17 cases were multiple tumors at first operation .The maximal tumor diameter was ≥ 3 cm in 13 cases and <3 cm in 28 cases . The first treatment was transurethral resection of bladder tumor(TURB‐t) .The pathological report was the stage T1 of urothelium cancer .Results All 41 cases were completed the operation smoothly ,and no serious complication occurred .In the postoperative pathological examination ,7 cases(17 .07% ) had tumor residue or tumor recurrence ,among them ,3 case had residue f tumor base and 4 cases were new onset tumor;the pathological grade at Re‐TUR in 1 case was increased from G2 to G3 .The follow up lasted for 3―27 months(average 13 .2 months) ,9 cases relapsed ,3 cases (42 .86% ,3/7) were positive at Re‐TUR and 6 cases(17 .65% , 6/34) were negative at Re‐TUR .Conclusion Re‐TUR for treating stage T1 of non‐muscle invasive bladder cancer is safe and feasi‐ble ,its significance to pick out high‐risk patient for conducting further active treatment ,which may have certain effect for reducing the recurrence rate of non‐muscle invasive bladder cancer .

Objective:To study the effect of chemokines CCL20 and CCL22 combined with skin-induced Treg on survival time of grafted skin.Methods: Skin grafting mice were divided into four groups, three mice per group, namely Treg group, Treg+CCL20 group,Treg+CCL22 group and control group.C57BL/6 mice were used as donor and BALB/c as acceptor, and the Treg cells were isolated from the mice induced by skin allograft.After skin grafted,CCL20 and CCL22 were subcutaneous injection every day,which lasted for 10 day.Survival time of skin in each group were observed and recorded.The Treg colonzation experiments were performed as follows.We firstly isolated Treg with Magnetic cell sorting system( MACS) and then labled them with 99 Tcm.After that we intravenously injected them into the mice.3 hours later,the mice were sacrifced and the radioactivity of organs were detected by GC-2016γradioim-munoassay counter.Results:①After Treg treated the survival time of skin grafted in antigen-induced Treg group was signifiantly longer than control group,when treg were cooperated with CCL20 and CCL22,the skin grafted showed more longer survival time than Treg and control groups( P<0.001 ).②After injection of induced Treg, Treg in autologous and allogeneic skin grafts goups were mainly distributed in autologous and allogeneic skin,accounting for 60% and 98% respectively.When cooperated with CCL20 or CCL22,the Treg were mainly distributed in liver.Conclusion:Chemokines CCL20 and CCL22 synergistically improved the effects of skin antigen induced Treg on survival time of skin graft,which probably related with the Treg colonization into the liver.

Objective To compare the effect of transurethral resection of prostate (TURP) and TURP combine with transure-thral incision of the bladder neck(TUIBN) in the treatment of small size prostate hyperplasia .Methods From March 2002 to Octo-ber 2010 ,124 cases of small-size prostate hyperplasia patients were treated .All patients were randomized to undergo TURP or TURP with TUIBN .There were 62 patients in TURP group .Of which ,54 cases were followed up .There were 62 patients in TURP plus TUIBN group .Of which ,50 cases were followed up .The treatment effect of the two different surgical methods was evaluated by comparing international prostate symptom (IPSS) ,maximum flow rate(Qmax) and post voiding residual urine volume(PVR) et al .Results No statistical difference (P>0 .05)was found in age ,medical histories ,prostatic weight ,IPSS ,Qmax ,PVR between the two groups before operation .But significant difference(P<0 .01)was observed in IPSS ,Qmax ,PVR between the two groups after operation .There was no statistical difference(P>0 .05) in resected tissue weight and follow up between the two groups .The inci-dence rate of bladder neck contracture was 4% in TURP plus TUIBN group ,and it was significantly lower than that of TURP group(11% )(P< 0 .01) .Conclusion TURP plus TUIBN was better for the treatment of small size prostatic hyperplasia than TURP .

Objective To determine plasma soluble HLA-G (sHLA-G) levels in patients with systemic lupus erythematosus (SLE),and to analyze its association with the possibility of organs or systems involvement in lupus patients.Methods Plasma samples were collected from 96 SLE patients and 74 healthy controls,and sHLA-G levels were determined by enzyme-linked immunosorbent assay.The sHLA-G levels in SLE patients and healthy controls were compared with students't-test.The difiefence of clinical and seroimmunological data among the SLE patients was assessed by chi-square test or students't-test.A value of P<0.05 was considered to be significant.Results Plasma concentration of sHLA-G was significanto higher in SLE patients than that in healthy controls[(230±192)U/ml vs(118±38)U/ml,P=0.0001].No relationship between plasma sHLA-G levels and SLE disease activity index(SLEDAI)was found(r=0.157,P=0.141).However,the patients with increased levels of sHLA-G had more severe disease activity (11±5 vs 8±5,P=0.027) and more central nervous system (CNS) involvement (24.2% vs 4.8%,P=0.007) in comparison with patients with normal plasma levels of sHLA-G.Conclusion The increased production sHLA-G,paralleled with more severe disease activity and higher CNS involvement,indicates that sHLA-G may play an important role in the pathogenesis of SLE.

Objective To evaluate the clinical value of pulse indicator continuous cardiac output monitoring in treating acute kidney injury(AKI) due to sepsis.Methods Sixty-two patients with AKI due to sepsis in the Central Hospital of Zhumadian from August 2013 to August 2016 were rolled in and divided into control group (34 cases) and observation group (28 cases) according to whether adopted pulse indicator continuous cardiac output monitoring.Six hours and 24 hours after fluid resuscitatinn,resuscitation fluid volume,heart rate(HR),central venous pressure(CVP),mean arterial pressure(MAP),vasoactive drugs dose of the two groups were observed.Treatment times,daily filtration volume of continuous renal replacement therapy and independent urine volume,serum creatinine level,the survival rate after treating for seven days in the two groups were compared.Results There was no significant difference in the HR,central venous pressure,mean arterial pressure at six hours after fluid resuscitation between the two groups(P ＜ 0.05).At 24 hours after fluid resuscitation,there was no siguificant difference in the HR in the control group compared with that at six hours after fluid resuscitation (P ＜ 0.05),the CVP and MAP were higher than those at six hours after fluid resuscitation(P ＜ 0.05);the HR in the observation group was lower than that at six hours after fluid resuscitation (P ＜ 0.05),MAP was higher than that at six hours after fluid resuscitation (P ＜0.05),but there was no significant difference in the CVP in the control group compared with that at six hours after fluid resuscitation (P ＜ 0.05).The HR and CVP at 24 hours after fluid resuscitation in the observation group were lower than those in the control group(P ＜0.05),while there was no no significant difference in the MAP between the two groups(P ＜ 0.05).There was no significant difference in the fluid resuscitation volume,the dose of noradrenaline and dobutamine after treating for six hours between the two groups (P ＜ 0.05).Twenty four hours after fluid resuscitation resuscitation fluid volume and dobutamine dose in the observation group were significantly lower than those in the control group (P ＜ 0.05).The duration of renal replacement therapy,the daily filtration volume and the blood creatinine after treating for seven days in the observation group were significantly lower than those in the control group (P ＜ 0.05),but the independent urine volume after treating for seven days in the observation group was significantly higher than that in the control group (P ＜ 0.05).After treating for seven days,there were 28 cases survival,the survival rate was 82.4％ (28/34),while there were 25 cases survival,the survival rate was 89.3％ (25/28);the difference of survival rate between the two groups was not statistically significant (x2 =0.59,P ＜ 0.05).Conclusion In patients with AKI due to sepsis,pulse indicator continuous cardiac output monitoring can be used to optimize fluid management,and it can improved the renal function.

Objective To evaluate the effects of diagnostic and treatment manipulation of the urinary diseases on serum prostate specific antigen(PSA) levels .Methods 80 male patients were recruited from urology surgery department of Shenzhen Tranditional Chinese Medicine (TCM ) Hospital ,Which included 13 cases with digital rectal examination (DRE) ,10 cases with catheterization , 12 cases with rigid cystoscopy ,17 cases with prostate biopsy ,28 cases with transurethral resection of the prostate (TURP) .Blood samples of 80 patients were collected before diagnostic and treatment manipulation of the urinary diseases and 24 h ,3 d ,7 d ,14 d af‐ter that ,respectively .Then ,serum total prostate antigen(TPSA) and free prostate antigen (FPSA) was measured .Results There was no effects of DRE on serum TPSA and FPSA levels(P>0 .05) .On the contrary ,serum TPSA and FPSA levels increased sig‐nificantly in patients with catheterization and cystoscopy(P<0 .05) ,and the duration was longer(7-14 d) .Serum TPSA and FPSA levels increased significantly(P<0 .05)in patients with TURP and biopsy at the 24th hour after manipulation and it began to de‐crease on the third day .Also ,the serum TPSA and FPSA levels decreased to baseline after 14 days .Conclusion There′re no effects of DRE on serum TPSA and FPSA levels .However ,serum TPSA and FPSA levels increase differently in patients with catheteriza‐tion ,cystoscopy ,biopsy and TURP ,but the durations were different ,too .

Transcription factor is one of the key factors in the regulation of gene expression at the transcriptional level. It plays an important role in plant growth, active components biosynthesis and response to environmental change. This paper summarized the structure and classification of bHLH transcription factors and elaborated the research progress of bHLH transcription factors which regulate the active components in plants, such as flavonoids, alkaloids, and terpenoids. In addition, the possibility of increasing the concentration of active substances by bHLH in medicinal plants was assessed. The paper emphasized great significance of model plants and multidisciplinary research fields including modern genomics, transcriptomics, metabolomics and bioinformatics, providing the contribution to improve the discovery and function characterization of bHLH transcription factors. Accelerating the research in the mechanism of bHLH transcription factors on the regulation of active components biosynthesis will promote the development of breeding and variety improvement of Chinese medicinal materials, also ease the pressure of resources exhaustion of traditional Chinese medicine home and abroad.
Alkaloids ; biosynthesis ; Basic Helix-Loop-Helix Transcription Factors ; chemistry ; classification ; genetics ; metabolism ; Flavonoids ; biosynthesis ; Plants, Medicinal ; genetics ; metabolism ; Terpenes ; metabolism

The medicinal fungi, which are of great importance in traditional medicine, are facing the problems of wild resources scarcity and low concentration of bioactive compounds. Velvet family and LaeA global regulator play a vital role in secondary metabolism and developmental programs, which are found in a wide variety of fungi ranging from Chytridiomycota to Basidiomycota. This review elaborates the structures and functions between Velvet family and LaeA protein. The Velvet family which shares the Velvet protein domain, including VeA (Velvet), VelB (Velvet like B), VosA (viability of spores A) and VelC (Velvet like C), acts on the regulation function is secondary metabolism and developmental programs such as asexual and sexual development. Furthermore, the function is affected by environmental factors such as light and temperature. LaeA protein which owns S-adenosylmethionine-dependent methyltransferase domain, coordinately regulates development and secondary metabolism by regulating and modifying the Velvet proteins. The regulation of LaeA is mediated by light receptor proteins. Therefore, clarifying the mechanism of Velvet and LaeA proteins in medicinal fungi will pave the way for nurturing medicinal fungi and improving production of bioactive compounds.
Fungal Proteins ; metabolism ; Fungi ; chemistry ; Gene Expression Regulation, Fungal ; Genes, Regulator ; Protein Structure, Tertiary ; Secondary Metabolism ; Structure-Activity Relationship