Acute undifferentiated leukemia (AUL) is a clinically rare subtype of acute leukemia (AL). AUL lacks the serial specific antigen expressions, and the blasts have no morphological features of myeloid differentiation. Due to the rarity of AUL in clinic, there is no unified understanding of clinical and biological characteristics. It′s difficult to diagnose and choose the optimal treatment for AUL patients. And the prognosis of patients with AUL is poorer compared with other types of AL. Thanks to the big development of biotechnology in recent years, the related researchers have a better understanding of this rare disease. This article reviews the progress of diagnosis and treatment in AUL.

ObjectiveTo optimize the condition of multiple fluorescence quantitative PCR and establish a new assay of four human herpes virus (HHV) detected by AllGlo quadruple fluorescence quantitative PCR.MethodsFour HHV including HSV-1,HSV-2,EBV and CMV were identified by sequence analysing the qualitative PCR production.Furthermore,they were quantitatively detected by AllGlo and TaqMan multiple fluorescence quantitative PCR respectively.ResultsBoth the positive rate and specificity of AllGlo and TaqMan in detecting single HHV achieved 100％.And AllGlo single fluorescence quantitative PCR prevailed over TaqMan's by Ct of 1-3.Four HHV can be simultaneously detected by AllGlo quadruple fluorescence quantitative PCR,comparing to the only two by TaqMan.ConclusionAllGlo fluorescence quantitative PCR assay allows a higher throughput,sensitivity and specificity than TaqMan in detection and thus provides a board prospect.

Objective:To investigate the effects of IRF 3 shRNA adenovirus on dynamic changes of early cytokines in LPS-stimulated primary Kupffer cells ( KCs ).Methods: Rat KCs were isolated and purified by means of in situ perfusion.After being infected with adenovirus carrying IRF 3 shRNA for 48 h, KCs were stimulated with LPS.Cell culture supernatants were collected respectively at 0,2,4 and 6 h after LPS stimulation as well as cells at 6 h.Supernatant cytokine secretion levels were detected by enzyme-linked immunosorbent assay ( ELISA).Intracellular gene expressions were tested by RT-PCR and Westeron blot.Results:IRF3 mRNA and protein were induced by LPS ,but suppressed by IRF 3 shRNA adenovirus in LPS-stimulated or non-stimulated KCs.IFN-βsecretions rose in the very early stage ( at 2 h) ,reached the peak at 4 h,and began to reduce but still remained high levels at 6 h after LPS stimulation in KCs.Interference adenovirus pretreatment suppressed IFN-βsecretions ( especially the secretion peak ) at each time point after LPS stimulation.IFN-βsecretions reached normal levels at 6 h after the stimulation in adenovirus-pretreated cells;TNF-αse-cretions rapidly increased in the very early stage and reached the peak at 2 h,then began to decrease gradually ,but remained high levels at 6 h after LPS stimulation in KCs.Interference adenovirus pretreatment inhibited LPS-induced TNF-αsecretions, especially the secretion peak;IL-1βsecretions did not increase untill 4 h, but reached a higher level at 6 h after LPS stimulation.Interference adenovirus suppressed IL-1βsecretions in the early stage of LPS stimulation;IL-10 secretions began to rise in the very early stage ,and gradually increased over time after LPS stimulation in KCs.Pretreatment of adenovirus with IRF 3 shRNA promoted upregulations of IL-10 secretions at each time point of the early of LPS stimulation.Conclusion:IRF3 gene expression can be silenced by IRF 3 shRNA ad-enovirus.IRF3 can promote its downstream signaling molecule IFN-βand pro-inflammatory cytokines including TNF-αand IL-1β,and block anti-inflammation cytokine IL-10 secretions in LPS-stimulated primary KCs.Therefore,IRF3 may play a central role in immune inflammatory injury of liver tissues.

Objective To investigate the impacts of adenovirus on the transfection efficiency and proliferative activity of primary Kupffer cells (KCs). Methods Rat liver KCs were separated and purified by density gradient centrifugation , and was then transfected with adenovirus carrying green fluorescence protein (GFP) gene at different multiplicity of infection (MOI). After 24 h, the transfection efficiency was evaluated by fluorescence microscope and flow cytometry. The proliferative activity of KCs was assessed by colorimetric method. Results The positive percentages of GFP staining cells were statistically different among different doses of adenovirus (MOI 0, 100, 300, 500, 700 and 900) under fluorescence microscopy or by flow cytometry (P <0.05 for all comparisons). The cell proliferative activity had significant differences among MOI 300, 500, 700 and 900(P < 0.05 for all comparisons), but had no differences among MOI 0, 100 and 300 (P > 0.05 for all comparisons) by CCK8 assay. Conclusions KCs can effectively be transfected by GFP adenovirus; and with an increase in virus MOI, the transfection efficiency rises gradually. A higher dose of adenovirus may have a negative effect on cell proliferative.

Objective:To investigate the changes of gene expression profile in lipopolysaccharide (LPS)-stimulated primary Kupffer cells ( KCs ) . Methods: Rat KCs were isolated and purified by means of in situ perfusion and density gradient centrifugation. After being identified by ink phagocytosis and ED2 staining test,KCs were stimulated with LPS. Gene expression profile were studied using gene microarrays,and the most significant upregulated gene was verified using real-time PCR. Results:27 genes were upregulated including Ces1f, Slc17a3, Slc21a4, Hsd17b2, Sorbs2, Ccdc116, Mgam, Myo5b, Etl4, Fabp1, Kif4b, Fosl1, Cyp4a1, Penk, Tmem221,Rpl5,Nr2f1,Hoxb1,Gpr165,Fam90a13p,Kpna6,Irak1bp1,Kcnh1 and 4 unnamed genes and 4 downregulated including Oc90,Tagln,Arxes2 and Olr830 in LPS-stimulated KCs. Among the upregulated genes, Ces1f was the most significant upregulatory gene. Real-time PCR confirmed that the levels of Ces1f were 23. 88 times higher in LPS-stimulated than control cells. Conclusion:There is a significant difference between LPS-stimulated and normal control cells in gene expression profile by microarray analysis,and Ces1f is the most significantly upregulated gene.

Objective To investigate the effects of interferon regulator factor 3 (IRF3) shRNA on the expression of TLR4 downstream signal molecules including IRF3-IFN-β, NF-κB/p38 MAPK-TNF-α/IL-1βand IL-10 in lipopolysaccharide (LPS)-stimulated Kupffer cells (KCs). Methods KCs were isolated from rats by in situ perfusion. The adenovirus strains carrying IRF3 shRNA were used for the transfection of purified KCs. The isolated KCs were randomly divided into four groups including adenovirus(-) LPS(-) treatment group, adenovirus(-) LPS(+) treatment group, adenovirus(+) LPS(-) treatment group and ad-enovirus(+) LPS(+) treatment group. The levels of cytokines in the supernatants of KC culture were detec-ted by enzyme-linked immunosorbent assay ( ELISA ) . Real-time PCR and Western blot assay were per-formed to analyze the expression of related cytokines at mRNA and protein levels, respectively. Results The expression of IRF3 at mRNA and protein levels in primary cultured KCs were induced by LPS. The cel-lular constitutive expression of IRF3 at mRNA level and the LPS-induced expression of IRF3 were signifi-cantly inhibited after transfection of KCs with adenovirus strains carrying IRF3 shRNA. However, the nucle-ar constitutive expression of IRF3 protein was not affected by IRF3 shRNA. The expression of IFN-βat mR-NA and protein levels in KCs were induced by LPS, but were suppressed by the interference with IRF3 shR-NA. No significant changes of the cellular constitutive expression of IFN-βat mRNA and protein levels were observed in IRF3 shRNA-treated KCs. Enhanced expression of proinflammatory cytokines including TNF-αand IL-1β at mRNA and protein levels were detected in LPS-stimulated KCs. Transfection of KCs with ade-novirus strains carrying IRF3 shRNA inhibited the LPS-induced secretion of TNF-α and IL-1β, but neither LPS-induced expression of TNF-α and IL-1β at mRNA level nor cellular constitutive expression of TNF-αand IL-1βat mRNA and protein levels were affected by IRF3 shRNA. The LPS-induced expression of IL-10 at mRNA and protein levels were enhanced in IRF3 shRNA-treated KCs. However, the cellular constitutive expression of IL-10 at mRNA and protein levels were not affected by the adenovirus. The levels of phosphor-NF-κB p65 subunit and phosphor-p38 MAPK protein in the nuclei of KCs were increased upon the stimula-tion with LPS. Treatment of KCs with IRF3 shRNA showed no significant effects on nuclear phosphor-NF-κB p65 subunit and phosphor-p38 MAPK. Conclusion Transfection of LPS-stimulated primary KCs with ade-novirus strains carrying IRF3 shRNA could effectively inhibit the expression of IRF3 and the transduction of downstream signals. IRF3 enhanced the secretion of TNF-αand IL-1β, but inhibited the expression of IL-10 in LPS-treated KCs. The LPS-induced activation of NF-κB and p38 MAPK in KCs were not affected by IRF3 signal.

Objective To investigate the trend of antimicrobial resistance and the prevalence of resistant genes in Pseudomonas aeruginosa strains isolated from Hangzhou First People's Hospital.Methods Antimicrobial susceptibilities of 1489 Pseudomonas aeruginosa strains isolated from 2003 to 2007 were statistically analyzed using WHONET.MICs of 11 antimicrobisis to 36 multi-drug-resistant Pseudomonas aerugionosa strains were determined by agar dilution method.Genes of β-lactamases(BLA)and aminoglycoside-modifying enzymes(AMEs)were detected by PCR and the PCR products were sequenced.Results The resistant rates to aztreonam,imipenem,ceftazidime,cefepime,piperacillin,piperacillin/tazobactam.cefoperazone/sulbactam,ciprofloxacin,levofloxacin,gentamicin and amikacin were increased from 13.4%,10.6%,8.7%,7.9%,12.7%,12.7%,6.7%, 15.8%,20.5%,24.7% and 10.9%in 2003 to 35.3%,40.9%,18.4%, 32.4%,32.9%,32.0%,21.9%,37.8%,38.6%, 39.4% and 34.8% in 2007.respectively.Hish MICs of 11 antimicrobiMs for multi-drug resistant Pseudomonas aerouginosa were determined with MIC90≥128 μg/mL.In 36 multi-drug resistant Pseudomonas aeruginosa strains,21(58.3%)strains carried β-lactamase genes and 32 strains(88.9%)carried aminoglycosidemodifying enzyme genes,while the deletion rate of oprD2 was 80.6%(29/36).Conclusions The resistant rates to common antibiotics of Pseudomonas aeruginosa have increased.resulting in multi-drug resistance.Genes of β-lactamases and aminoglycoside-medifying enzymes are prevalent in multi-drug resistant Pseudomonas aeruginosa strains,with the common deletion of oprD2.

Objective: To investigate the effect and safety of interleukin-2 combined with cisplatin in the treatment of malignant pleural effusion.Methods: Totally 86 patients with malignant pleural effusion were randomly divided into the observation group (44 cases) and the control group (42 cases).The control group was treated with cisplatin, and the observation group was treated with interleukin-2 additionally.The treatment course was 3 weeks.The ascites relief, quality of life, survival status and tumor markers before and after the treatment, and the adverse drug reactions were compared between the groups.Results: The total ascites remission rate and the effective rate in the observation group were significantly higher than those in the control group (P<0.05).Compared with those before the treatment, CEA, CA19-9 and CA450 decreased significantly after the treatment in both groups (P<0.05), those in the observation group were significantly lower than those in the control group (P<0.05).After the treatment, the physiological function, physiological occupation, emotional function, body pain and the other life quality scores in the observation group were significantly higher than those in the control group (P<0.05).The 24-month survival rate in the observation group was significantly higher than that in the control group (P<0.05).There were no significant differences in adverse drug reactions between the groups (P>0.05).Conclusion: Interleukin-2 combined with cisplatin can effectively relieve ascites and improve quality of life, which can also prolong the survival time of patients.

Objective To evaluate hepatitis B virus large surfsce protein(LHBs) in monitoring of antiviral treatment.Methods LHBs.HBV serum markers and HBV DNA loads in 46 patients with adefovir dipivoxil(ADV) therapy were monitored for the whole course(60 weeks).Enzyme linked immunosorbent assay(ELISA),time differentiate immunofluoresence assay and real.time polymerase chain reaction(RT-PCR)were performed to detect LHBs,HBV serum markers and HBV DNA loads,respectively.And correlation analysis was also performed.Results Both LHBs and HBV DNA declined during the ADV treatment.and the correlation coefficient was 0.97.but LHBs declined after HBV DNA.There were 20 patients with HBV DNA<5×102/mL at 60th week.in which 8 were LHBs negative;in 14 recurrent patients,the HBV DNA turned to negative and became positive again,3 with negative LHBs;while in 12 patients resistant to the ADV therapy.2 were LHBs negative.Conclusion Dynamic monitoring of LHBs is useful in the evaluation of antiviral treatment.

Objecitve:To investigate effects of urotensin Ⅱ( UⅡ)/UT system on innate immune inflammatory signal pathway TLR4-IRF3 in the lipopolysaccharide (LPS)-stimulated Kupffer cells (KCs).Methods: Rat KCs were isolated and cultured.Pro-in-flammatory cytokines including IL-6,IFN-βand IFN-γwere assayed by ELISA in culture supernatant of KCs.Cell surface TLR4 were tested with flow cytometry technique.Expression of IRF3 were tested with real-time PCR and Western blot.Results: Significant increases were showed in IL-6, IFN-βand IFN-γsecretion, TLR4-expressed positive rates and IRF3 mRNA levels in KCs after stimulated by LPS,but were inhibited via urantide pretreatment.In addition,LPS induced upregulation of nuclear IRF3 protein and downregulation of cytoplasm IRF3 protein in KCs,which were blocked by urantide pretreatment.Conclusion:UⅡ/UT system mediates immune inflammatory response in part through activating TLR 4-IRF3 pathway in LPS-stimulated KCs.