Objective To explore the inhibitory effect of mutant influenza A viruses to the activation of interferon regulatory factor 3 (IRF-3). Methods HEK293 cells were infected with A/FM/1/47,A/HK/1/68, A/HK/1/68-MA20, A/HK/1/68-MA20C and positive control Sendai virus (SV). Whether the slowly moved phosphorylation form Ⅲ and Ⅳ of IRF-3 appeared or not was compared by Western blot in cells infected with these viruses. Wild type of NS1 from A/HK/1/68 and mutant NS1 from A/HK/1/68-MA20 were subcloned into pcDNA3.1-flag respectively. They were transfected in HEK 293 cells respectively. At 16 hours posttransfection, cells were infected with Sendai virus for 8 hours. Whole cell extracts were analyzed by Western blot and then probed with monoclonal flag antibody to check the expression of NS1, or with anti-IRF-3 to observe the inhibitory effects of the wild and mutant NS1 to the activated IRF-3. Luciferase assay was carried out by co-transfection with reporter plasmid, pGL2B with interferon ? promoter, and wild or mutant NS1 cDNA expression plasmid. SV was used to infect these cells after the co-transfection. Results Only less virulent A/HK/1/68-MA20 and positive control SV can activate IRF-3. Activated form Ⅲ and Ⅳ of IRF-3 began to appear 9 hours post infection (h.p.i), and most significant activated IRF-3 appeared 23 and 26 h.p.i. Sequence analysis of NS1 of MA20 revealed that nucleotide position number 94 is mutated from T to C, and amino acid at position number 23 is changed from valine to alanine. Co-transfected with wild type NS1 made form Ⅲ and Ⅳ of IRF-3 almost disappear, but not mutant NS1. In the luciferase functional analysis, wild type NS1 can inhibit the luciferase activity of IFN-? promoter, which was induced by SV, to around 1/10. Again no inhibitory effects was observed of mutant NS1 in the luciferase assay. Conclusion The mechanism that A/HK/1/68-MA20 can activate IRF-3 is that point mutant NS1 abolished the inhibitory function of NS1.

Objective Interferon regulatory factors 3 (IRF-3) is a key transcription factor to regulate gene expression of interferon after virus infection. This study aims to look for new spliced isoforms of IRF-3 and to investigate their structures and functions. Methods RNA extracts from human embryonic kidney 293 cells were amplified by RACE and RT-PCR. New sequences were compared with published sequences of IRF-3 and murine EST database using bioinformatics method. A new sequence, IRF-3c, was subcloned into pcDNA3.1-flag. The IRF-3c/pcDNA3.1-flag plasmid was transfected in HEK 293 cells. Whole cell extract was analysed by Western blot and then probed with monoclonal Flag antibody. Luciferase assay was carried out by co-transfection with reporter plasmid, pGL2B with interferon ? promoter, and IRF-3c cDNA expression plasmid. At 16 hours posttransfection, cells were infected with Sendai virus for 8 hours. Cells were collected and assayed for luciferase activity. Results A novel spliced isoform of IRF-3, named IRF-3c was discovered. The new isoform is almost the same as IRF-3, except for the utilization of the 180 bp bases in intron 6 adjacent to exon 6. The first 2,3 and 4 bases are a stop codon, which may produce a protein with a truncated C-terminal stoped at amino acids 327. Western blot analysis confirmed an expected 44 kDa strong band. The new inserted bases can be found in murine EST database, suggesting a conservative function in evolution. The functional luciferase assay showed that IRF-3c inhibited the IFN? promoter activity to (around) 40%～50% as that of control after Sendai virus infection. Conclusions The discovery of a new isoform of IRF-3 provides a new insight into the functional regulation of IRF-3 family. It is a dominant-negative inhibitor for interferon ? promoter activity in the virus infection pathway, provides a mechanism for the fine-tuning of the virus-induced activation of the interferon response, and prevents interferon ? from its overexpression and its toxic effects. It is worthwhile to explore the role of IRF-3c in the pathogenesis of human diseases using IRF-3c’s specific sequence.

AIM: To investigate the effect of IL-10 on IL-1?-induced prostaglandin E 2(PGE 2) release and cyclooxygenase-2(COX-2) expression in human mesangial cells and to examine whether IL-10 has effect on the biological function of IL-1?.METHODS: The PGE 2 concentration in supernatants of HMC was measured by radioimmunoassay. The COX-2 mRNA and protein were measured by RT-PCR and Western blot, respectively. RESULTS: PGE 2 and COX-2 were significantly increased after treatment with IL-1?( P0.05 , respectively), while it inhibited IL-1?-elicited PGE 2 production, as well as COX-2 mRNA and protein expression in a concentration-dependent fashion. CONCLUSIONS: These results indicated that IL-10 depressed the IL-1?-induced release of PGE 2 and expression of COX-2. These data suggested that IL-10 could exert anti-inflammatory actions at several levels, not only by inhibiting the production of pro-inflammatory cytokines but also by suppressing their biological function.

Objective To evaluate the efficacy and safety of low-dose dexamethasone pretreatment regimen in prevention of hypersensitive reaction (HR) related to docetaxel in elderly tumor patients. Methods According to the order for admission and the ratio of 3:2, 91 elderly patients with docetaxel weekly therapy were randomly divided into two groups: experimental group and control group. All patients aged from 65 to 82 years with a median age of 68 years old. There were 54 patients in the experimental group and 37 patients in the control group. In the experimental group, patients received oral dexamethasone 4. 5 mg once daily on 1 day before treatment, the day of treatment and continuing for 3 days after treatment, while patients received 8 mg twice daily in the control group. All patients were scored according to MCIRS by the physician. The side effects were evaluated by NCI-CTCAE3.0. Results Four cases in the experimental group (7.4 %) and three cases in the control group (8. 1%) occurred HR, and there was no significant statistical difference (P=1. 000). Conclusions The low dose dexamethasone is efficient and safe compared with the conventional dose dexamethasone, and there is no significant difference in HR incidence between two groups.

ObjectiveTo explore the clinical manifestations of infant cytomegalovirus infection in different age. MethodThe clinical data of 237 infants who suffered from cytomegalovirus infection was analyzed retrospectively and divided into three groups: 0-3 months old (77 cases,group A) ,4-6 months old ( 65 cases , group B ) , 7-12 months old ( 95 cases , group C ). ResultsThe incidence of respiratory infection was the highest among all infectious organs in three groups, the numbers of patients who had wheeze in group A was less than that in the other two groups[24.7％(19/77) vs. 61.5％ (40/65), 61.1％( 58/95 )](P ＜ 0.01 ).The incidence of jaundice decreased gradually as the babies grew up[23.4％( 18/77 )→7.7％(5/65 )→1.1％( 1/95 )](P ＜ 0.05). There was no significant difference in diarrhoea and bleeding among three groups (P＞0.05 ). The proportion of alanine aminotransferase], aspartate amino transferase increasing was similar among three groups, but gamma-glutamyl transferase (GGT) was different, the proportion of GGT increasing was 77.9％ (60/77) in group A which was higher than that in the other two groups[10.8％ ( 7/65 ), 2.1％ ( 2/95 )].Granulocytopenia in group B was obviously decreased compared with the other two groups (P ＜ 0.05 ), anemia was easily occurred in group C (P ＜ 0.05 ). ConclusionThe injury of cytomegalovirus infection may be related to month old.

Objective We investigated the value of endoscopic evacuation and craniotomy of the hypertensive in?tracerebral hemorrhage to determine which methods are more suitable for the patients. Methods One hundred twenty pa?tients with hypertensive intracerebral hemorrhage participated this study. They were divided into classic surgical evalua?tion group (n=60) and endoscopic surgical evaluation group (n=60) according to their corresponding surgery strategies. Each patient was assessed by the preoperative Glasgow Coma Scale (GCS), the mean rate and time of hematoma evacua?tion from onset to operation, the postoperative GCS, the mean time of admission in neuro-intensive care unit (NICU) and Glasgow Outcome Score (GOS) at 3 month after surgery. Results The continuous (≥3 months) follow-up surveys were all completed by 120 patients. There was no statistical difference in clinical data before operation between two groups (P>0.05). However, clearance of hematoma was much faster and more efficient in endoscopic surgical group than in classic surgical evaluation group (1.5 ± 0.4 vs.3.9 ± 0.6 h, P<0.01; 95.84 ± 2.72% vs.87.48 ± 7.84%, P<0.01). The GCS scores were 10(6,12),12(8,13) and 13(10,13) in endoscopic surgical group whereas were 6(5,9),7(5,11).8(5, 12) in craniotomy group at 1,3 and 7 d followed operation. GCS scores were higher in surgical group than in craniotomy group at all time points (P<0.01). In addition, patients receiving endoscopic treatment showed a shorter NICU admission time than those receiving craniotomy (3.55±4.21d vs. 9.10±4.72d, P<0.01). The intracranial infection and hypostatic pneumonia were sig?nificantly lower in endoscopic than in craniotomy surgery group (0 vs.6 cases; 5 vs. 41 cases, P<0.05). The endoscopic treatment significantly improved the GOS score compared with craniotomy [3(3, 4)vs. 2(2, 3)] (P<0.01). Conclusion Endoscopic evacuation of hematoma for hypertensive intracerebral hemorrhage is efficient and minimally invasive, which is superior to craniotomy.

Purpose To explore the clinical value of color Doppler ultrasound in monitoring hemodynamic changes in main renal artery of neonatal asphyxia. Materials and Methods A total of 60 cases of neonatal asphyxia were divided into mild asphyxia (38 cases) and severe asphyxia (22 cases) according to Apgar score 1 min after born. Then the peak systolic velocity (Vs), the end diastolic velocity (Vd) and the resistance index (RI) of the main renal artery were obtained by color Doppler ultrasound on day 1 and day 3;the level of endothelin-1 (ET-1) was also recorded accordingly. The above results were compared with those of 20 cases of healthy full-term new born infants. Results On day 1, Vs and Vd of the main renal artery in the groups with mild asphyxia and severe asphyxia were both lower than those in healthy group (P<0.05), but RI was higher (P<0.05), with more dramatic changes in the group with severe asphyxia (P<0.05). On day 3, Vs and Vd in the groups with mild asphyxia and severe asphyxia reduced compared with those on day 1, whilst RI was higher than that on day 1. Vd and RI in the group with severe asphyxia changed more significantly (P<0.01). As to the value of ET-1, both groups with mild asphyxia and severe asphyxia showed higher level than healthy group (P<0.01). More dramatic increase appeared in the group with severe asphyxia (P<0.05). In the groups with mild asphyxia and severe asphyxia, the Vs and Vd of the main renal artery were negatively correlated with ET-1 on day 1 and day 3 (r=-0.823,-0.845;P<0.01), while the RI was positively correlated with ET-1 (r=0.785, P<0.01). Conclusion Both color Doppler ultrasound imaging and neonatal urine ET-1 test can reflect degree of renal injury after neonatal asphyxia dynamically and noninvasively, which can be used to evaluate the injury severity.

Objective To investigate the number, structure and demand of rehabilitation professionals hospitals above grade 2A in Jiangsu. Methods 176 hospitals above grade 2A in Jiangsu were investigated with questionnaire in the topics of general condition of department of rehabilitation medicine, structure of rehabilitation professionals and demand of rehabilitation professional. Results 64% of the hospitals has established department of rehabilitation. Rehabilitation professionals, such as the rehabilitation physician, therapists, nurses and engineer were in a total of 1657, and 231 professionals were needed further. Conclusion There are problems of lack of the department of rehabilitation medicine and rehabilitation professional, with the imbalance structure of professional in hospitals above grade 2A in Jiangsu.

Objective To identify the important regulatory elements in the promoter of human CD2 associated protein(CD2AP) by conserved sequence analysis among different species and luciferase functional detection. Methods The promoter sequences of CD2AP from different species were analyzed by BLAST. Plasmids containing different length of deletion mutations of human CD2AP promoter were constructed. Pro-moter activities were tested in 3 kinds of cells from different species by luciferase analysis and were tested in HEK-293 cells treated with all-trans-retinoic acid. Results Homologous sequence comparison in CD2AP promoter area among human, cattle and pig showed that putative specific protein 1 (Sp1) sites and down-stream promoter element (DPE) were highly evolutional]y conserved. Progressive deletion luciferase analysis of DNA fragments revealed similar promoter activity style among 3 different cell lines from 3 different spe-cies, HEK-293, BHK-21 and Vero cells. One basic promoter activity located within 500 bp upstream of ATG. Fragments of further upstream 100 bp or more had drastically 10 times increased promoter activity. Two putative Sp1 sites were in this 100 bp region. All-trans-retinoic acid decreased the luciferase activity of CD2AP promoter. Conclusion Putative Spl sites and DPE have important functions in the promoter activity of CD2AP.

Purpose To investigate the effect of small interfering RNA on the increase of myocyte enhancer factor 2A(MEF2A) expression in PC12 cells exposed to hypoxia.Methods PC12 cells were cultured under normal conditions or were exposed to hypoxic conditions.Small interfering RNA targeted MEF2A gene (MEF2A-siRNA) was chemically synthesized. Eukaryocytic expression vector was built and transfected into PC12 cells with liposome. The expression of MEF2A mRNA was detected by real-time PCR. Western blot was used to detect the MEF2A protein.Results Compared with normal control(2~(-△△CT)=1.01±0.02), the mRNA level of MEF2A gene in PC12 cells with the treatment of MEF2A-siRNA was down-regulated significantly by 88%(2~(-△△CT)=0.12±0.03, P＜0.01).The expression of MEF2A protein in hypoxia-treated PC12 cells was markedly higher than that of normal control(98.4±11.7 and 47.5±7.6,P＜0.01).However, MEF2A-siRNA could significantly suppress the increase of MEF2A protein (P＜0.01).Conclusion MEF2A gene silence induced by siRNA might inhibit the increase of MEF2A protein by hypoxia in PC12 cells.