Objective: To establish a method for the determination of 17 heavy metals and harmful elements in Xiyanping Injection (XI). Methods: The samples were digested and determined by inductively coupled plasma mass spectrometry (ICP-MS). The RF power was 1 550 W; The peristaltic pump speed was 0.3 r/s; The plasma gas velocity was 15 L/min; The auxiliary gas velocity was 0.2 L/min; The carrier gas velocity was 1 L/min; The sampling depth was 10 mm and the signal was measured for three times. Results: The detection limits of Li, Al, V, Cr, Fe, Co, Ni, Cu, As, Ag, Cd, Sn, Sb, Ba, Hg, Pb, Bi were 9.584, 49.858, 0.504, 3.016, 51.209, 0.142, 1.116, 0.675, 0.924, 1.421, 0.403, 2.770, 0.711, 3.584, 0.590, 0.411, 0.169 ng/mL, and the quantitative limits were 28.933, 151.085, 1.528, 9.139, 155.179, 0.429, 3.381, 2.046, 2.799, 4.312, 1.220, 8.394, 2.155, 10.861, 1.965, 1.244, and 0.513 ng/mL within a certain concentration range, respectively. The response value had a good linear relationship with the concentration (r > 0.999 1); The RSDs (n = 12) of the injection precision test were in the range of 0.8%-3.8%, the RSDs (n = 6) of the repeatability were in the range of 0.7%-2.0%, and the average recoveries of 17 elements were in the range of 95.7%-104.8%. Detection results of 20 batches of mass production samples: Ag was not detected, Li and V were detected but lower than the quantitative limit, the maximum mass concentration of Al, Ni, Cu, As, Ba, Hg and Pb were 0.685, 0.013, 0.007, 0.006, 0.208, 0.070 and 0.027 μg/ml respectively, Cr, Fe, Co, Cd, Sn, Sb, and Bi were not detected or lower than the quantitative limit. Conclusion: The specificity, linearity, sensitivity, precision, accuracy and durability of the method were all good. It can be used to determine 17 elements of Li, Al, V, Cr, Fe, Co, Ni, Cu, As, Ag, Cd, Sn, Sb, Ba, Hg, Pb and Bi in XI. The test results of 17 elements in 20 batches of mass production samples were less than the limit requirements, meeting the requirements.

Objectives: To observe the drug seeking behavior in chronically morphine pretreated, abstinent rats and determine brain regions involved in this behavior. Methods: Preferences for morphine associated environments and Fos expression in some brain regions were measured 35 days after withdrawal of chronic morphine treatment. Results: The preference for the morphine environment was greatly increased by morphine pretreatment. The conditioned morphine pretreated group showed significantly higher Fos levels than either the conditioned or non conditioned control groups in the Cg, Ac C, BNST VL, ACE and ABL. Conclusions: A relationship between place preference behavior and neural indices of activation exist in the forebrain in response to morphine conditioned cues, which may be chronically modulated by prior morphine exposure.

Objective To investigate the effect of microRNA-21 (miR-21) antisense oligonucleoti-de on collagen synthesis in the rat hepatic satellite cells ( HSCs) .Methods Rat hepatic stellate cells were isolated and cultured; the miR-21 antisense oligonucleotide was transfected into HSCs with lipofectamine 2000;after incubation 48 h, the HSCs were collected.The expression of miR-21 was detected with reverse transcription polymerase chain reaction ( RT-PCR) , andα-smooth muscle actin (α-SMA) protein in HSCs with Western blot.The cell proliferation was assayed with methyl thiazolyl tetrazolium ( MTT) method.Re-sults Compared to scrambled control group, the expression of miR-21 was reduced by 76%( P <0.01), the proliferation activity of HSCs was reduced by(26 ±3)%( P <0.01), the expressions of type I and III collagen proteins were reduced by(61 ±7)%and (48 ±6)%( P <0.01).Conclusions The miR-21 an-tisense oligonucleotide could reduce significantly the expression of miR-21, and inhibit HSC proliferation and extracellular matrix production.

Objective To investigate the distribution of five indicators of hepatitis B(HBV-M)and its relationship with liver function parameters and HBV-DNA load in patients with chronic HBV infection in different stages.Methods The serum samples were collected from 456 patients infected with HBV.The HBV-M,liver functional parameters and HBV-DNA level were quantita-tively detected.According to the stages of disease,the patients were divided into 3 groups including chronic hepatitis B group(inclu-ding mild subgroup,moderate subgroup and severe subgroup),liver cirrhosis group(including compensatory subgroup,decompen-sated subgroup)and hepatocellular carcinoma goup.Results The ratio of HBsAg,HBeAg,HBcAb positive pattern(135 pattern) and HBsAg,HBeAb,HBcAb positive pattern(145 pattern)in the three groups were statistically different(P <0.05).In each chron-ic hepatitis B subgroup,both ALT and AST levels of 135 pattern were significantly higher than the other two patterns(P <0.05). In each liver cirrhosis subgroup and hepatocellular carcinoma group,there were statistically significant differences in ALB and TBIL levels between the three patterns(P <0.05 ).In each group,the HBV-DNA level of 135 positive pattern were significantly higher than the other two patterns(P <0.05).Conclusion With the advancement of chronic hepatitis B,there is a downtrend in the ratio of 135 pattern and increasing trend in the ratio of 145 pattern.If the stage of hepatitis B is discriminated,ALT,AST,ALB TBIL and HBV-DNA level were closely related to HBV-M pattern.

Background Infectious keratitis is common blinding eye disease in China.Inflammatory runaway reaction often occurs in infectious keratitis, and its mechanism and treating approach are worthy of research.Objective This study was to investigate the clinical manifestation,mechanism and treatment outcome of doxycycline in overwhelming inflammation induced by Pseudomonas aeruginosa keratitis.Methods Pseudomonas aeruginosa suspension was intrastromal injected in 60 right eyes of 60 New Zealand white rabbits,and gatifloxacin eye drops was frequent instilled for consecutive 3 days to establish corneal inflammatory runaway reaction models,and 42 eyes with worse keratitis were defined as overwhelming inflammation.Then the models were randomly divided into gatifloxacin treatment group (15 rabbits), combined treatment group (gatifloxacin with doxycycline treatment, 15 rabbits) and balance salt solution (BSS) control group (12 rabbits), and corresponding eye drops was topically administered in the rabbits 8 times per day for 14 days.The symptoms of the models were observed under the slit lamp microscope on day 1,3,7 and 14 after treatment,and the corneal infiltration area was calculated.The corneal morphology was examined by using hematoxylin-eosin staining.The expressions of matrix metalloproteinase-2 (MMP-2) and MMP-9 in the corneal tissue were examined by immunohistochemistry.The contents of tumor necrosis factor α (TNF-α) and interleukin-1β (IL-1β) in corneal homogenate were detected by ELISA.The use and care of the animals complied with the ARVO Statement.Results The inflammatory scores were lower and infiltration area of cornea was smaller 7 days after treatment than those before treatment in the combined treatment group and gatifloxacin treatment group (both at P＜0.05).Hematoxylin-eosin staining result showed that the inflammation response of cornea subsided 14 days in comparison with 5 days after treatment in the combined treatment group, with the proliferation and rearrangement of fiberal tissue.The MMP-2 and MMP-9 were strongly expressed in corneal tissue with overwhelming inflammation in day 5 after treatment, however, the expression intensity weakened in 14 days after treatment in the combined treatment,showing significant differences in the absorbance between them (all at P＜0.05).ELISA assay showed that the contents of TNF-α and IL-1β in corneal tissue were significantly lower in the combined treatment group than those in the BSS group (P =0.00,0.03).In addition, contents of TNF-α and IL-1β in corneal tissue were significantly declined at day 14 in comparison with day 5 and day 7 after treatment in the combined treatment group (all at P＜0.05).Conclusions Overwhelming inflammatory keratitis models can be successfully established by corneal intrastromal injection of Pseudomonas aeruginosa suspension and immediately frequent 3-day instillation of gatifloxacin eye drops in rabbits.The topical application of doxycycline plays treating effect on overwhelming inflammatory keratitis by inhibiting the release of pro-inflammatory cytokines and MMPs.

A novel antimicrobial peptide, named as perinerin (GenBank accession No. P84117), was isolated and characterized from Asian marine clamworms, Perinereis aibuhitensis Grube. Perinerin showes powerful and broad activity against both grampositive and gramnegtive bacteria in vitro, especially on Pseudemonas aeruginosa. To obtain large amounts of active perinerin and characterize its main physiochemical features, The perinerin weve expressed in Pichia pastoris. Intact perinerin gene amplified by the modified gene SOEing method(Gene splicing by overlap extension)was cloned into expression vector pPICZ?A and obtained recombinant vector pPICZ?APEN, then pPICZ?APEN was expressed in the Pichia pastoris GS115. The expressed sample was analyzed by TricineSDSPAGE. The results showed that Pichia pastoris was a suitable system producing the secreted form of perinerin. Bioactivity assay showed that the recombinant perinerin had marked antimicrobial effects.

Background The study on the classification of fungi is very important for the diagnosis and treatment of fungal keratitis.Identifying the different species of filamentous fungi is a critical factor for the application of anti-fungal drug in treating keratitis.ObjectiveThis report studies the relationship between the genotype of filamentous fungi and the clinical factors.MethodsFifty-two patients with filamentous fungal keratitis determined by clinical and laboratory examination were recruited in Tongren Hospital from January 2006-December 2006.The lesions were graded on the severity of the corneal ulcer and the presence of hypopyon.The filamentous fungal keratitis was treated with topical and systemic administration of anti-fungal drugs or corneal transplantation.The isolates were cultured in potato culture and identified by morphological characteristics based on the Nelson criterion and genotyped by the rDNA ITS method.The clinical data was retrospectively analyzed.ResultsForty-eight species (eubacteria are bacteria,not fungi)of fungus were identified by morphological characteristics,and the filamentous fungi were divided into 4 types based on the phylogenetic relationships within the rDNA ITS of the 52 filamentous fungi.The morphological characteristics and genotype were confirmed in 48 strains of eubacteria and 31 strains of 52 filamentous fungi (90.3%).The 4 groups of fungi were classified by genotype as follows:group 1 represents 22 strains including 20 strains of Fusarium solani and 2 strains of Fusarium oxysporum;group 2 represents 12 strains including 8 strains of Fusarium moniliformis,3 strains of Fusarium proliferatum and 1 strain of Fusarium incarnatum;group 3 represents 5 strains including 1 strain of Fusarium moniliformis and 4 unknown strains;group 4 represents 13 strains including 10 strains of Aspergillus spp.and 3 strains of Alternaria spp.Significant differences were found in the disease duration (P=0.00),inducing cause (P=0.03),ulcer grade (P=0.01)and outcome of the anti-fungal treatment (P=0.035)when compared between group 1 and 2 with group 3 and 4.Conclusion Filamentous fungi that cause keratitis could be correctly identified by sequencing the internal tanscribed spacer of rDNA.There are significant clinical differences among the groups classified by genotype.

Background Staphylococcus aureus,Staphylococcus epidermidis and Pseudomonas aeruginosa are common pathogenic bacteria in ophthalmology.To seek an optimal antimicrobial method is critical for the prevention of infection.Objective This study was to investigate the antimicrobial efficacy of riboflavin-ultraviolet A (UVA) on Staphylococcus aureus,Staphylococcus epidermidis and Pseudomonas aeruginosa.Methods Ten strains of frozen Staphylococcus aureus,Staphylococcus epidermidis and Pseudomonas aeruginosa were incubated to culture dishes and resuscitated for 2 times.The bacteria were put into bacterial dilution to prepare the suspension of 0.5 Mclntosh concentration and then the suspension was diluted 10-folds.UVA was used to irradiate the culture dishes added 0,0.1％,0.5％ riboflavin for 10,20,30 minutes with the exposure intensities of 1,2,3 mW/cm2,and the optimal bacteriostatic condition was evaluated by measuring the diameters of inhibition zone.Staphylococcus aureus,Staphylococcus epidermidis and Pseudomonas aeruginosa were divided into normal control group,riboflavin group,UVA group and photodynamic therapy (PDT) group (riboflavin+UVA),and colony forming unit was compared among the groups.Results The optional experimental conditions of PDT inactivating microorganisms were determined as UVA wavelength 370 nm,UVA exposure rate 3 mW/cn2,UVA exposure time 30 minutes and riboflavin 0.1％.The colony forming units were decreased in turn in the normal control group,riboflavin group,UVA group and PDT group (F =110.990,190.841,118.197,all at P =0.000).Compared with the normal control group,the colony forming units were significantly decreased in the UVA group (t =14.640,7.550,5.320,all at P＜0.05),and the colony forming units were significantly decreased in the PDT group (t =25.480,14.750,12.920,all at P＜0.05).The colony forming units decreased by 46.4％-50.2％ in the UVA group and 91.6％-96.7％ in the PDT group.There was no significant difference in colony forming units between the normal control group and the riboflavin group(t =0.660,0.470,0.510,all at P＞0.05).Conclusions PDT can effectively inactivate common bacteria in ophthalmology.This result suggests that PDT has a potential in the treatment of microbial keratitis.

Objective To study the diagnostic value of common inflammatory markers in patients with infectious diseases. Methods One hundred sepsis patients, 100 viral infection patients,100 pulmonary tuberculosis patients and 100 gonorrhea patients were analyzed retrospectively. The contents of procalcitonin (PCT), C-reactive protein (CRP), haptoglobin (HP), ceruloplasmin (CER), α1-acid glycoprotein (α1-AAG), α1-antitrypsin (α1-AAT), white blood cell count (WBC) and erythrocyte sedimetation rate (ESR) were measured. The receiver operating characteristic curve (ROC curve), sensitivity, specificity, positive predictive value, negative predictive value, Youden's index,positive and negative likelihood ratios and total coincidence rate were calculated respectively. Results The area under the ROC curve, sensitivity, specificity, Youden's index and positive likelihood ratios,positive predictive value and total coincidence rate of PCT in sepsis patients were 0. 895, 0.84, 0.92,0.76, 10.50, 0.91 and 0.88, respectively, which were superior to CRP, HP, CER, α1-AAG, α1-AAT, WBC and ESR. Conclusions PCT is a better inflammatory reactive parameter than other parameters currently applied in practice and may serve as a rapid and sensitive test in the early stage of severe bacterial infections.

Objective To investigate the effect of TGFβ1 siRNA on hepatic satellite cells (HSCs) activation, proliferation and extracellular matrix production. Methods The TGFβ1, siRNA plasmid was transfected into HSCs with Lipofectamine 2000. The supernatant and HSCs were collected after incubation for 72h. The expression of TGFβ1, and a-SMA protein in HSCs was detected by. Western blotting. The expression of type Ⅰ and Ⅲ collagen mRNA was detected by RT-PCR. The cell proliferation was assayed by MTT method. Contents of typeⅣ collagen and hyaluronic acid in supernatant were determined by radioimmuno-assay.Results Compared with scrambled control group, the TGFβ1, and a-SMA protein expression,the activity of HSCs proliferation,the expression of typeⅠ and Ⅲ collagen mRNA,and the contents of type Ⅳ collagen and hyaluronic acid in supernatant were reduced in TGFβ1, siRNA group by (79±5)%,(55±4)%, (25±4)% ,(63±6)% ,(57±4)% ,(53±8)% ,(46±8)% ( P<0.01),respectively.Conclusion TGFβ1, siRNA could significantly reduce the expression of TGFβ1,inhibited HSC activation,proliferation and extracellular matrix production.