Objective To clone and compare RNase HIIa and RNase HIIb of Chlamydia pneumoniae AR39(CpRNaseHIIa and CpRNaseHIIb). Methods Genes of CpRNase HIIs were amplified with the designed primers.Then,the recombinant plasmids were constructed,and CpRNase HIIs were expressed and purified with pET expression system.The 5′-32P-labeled RNA/DNA substrate and DNA with oligoribonucleotides substrates were prepared to identify characterization of CpRNase HIIs. Results Ribonuclease H activity of both CpRNase HIIa and CpRNase HIIb could cleave oligoribonucleotides strand,generating break with 3′-OH and 5′-phosphate ends.The results showed that there was different biochemical characterization between them. Conclusion CpRNase HIIa and CpRNase HIIb have different functions in C.pneumoniae.

[ Objective] To explore the variation features of contrast enhanced ultrasound ( CEUS) quantitative parameters and its correlation with tumor microvessel density ( MVD ) , after the treatment of living liver cancer using high intensity focused ultrasound ( HIFU ) . [ Methods ] A group of 30 New Zealand white rabbits were made into VX2 liver cancer models, that were randomly divided into control group ( n =15, not HIFU treatment) and treatment group ( n =15, HIFU treatment).CEUS was performed, then obtaining ascending slope(AS),arrival time(AT),time-to-peak(TTP),peak intensity ( PI) ,area under curve ( AUC ) .And correlation analysis was performed with MVD obtained by immune histochemical method. [ Results ] Compared with those in the control group, AS, PI, AUC were significantly decreased in treatment group (P <0.05) and so was MVD (P <0.05).AT,TTP were significantly delayed than those in the control group (P<0.05).The AS, PI, AUC of CEUS quantitative parameters were positively correlated with MVD (P<0.05).The AT, TTP respectively showed a negative correlation with MVD ( P<0.05) . [ Conclusion] CEUS quantitative parameters of liver cancer tissue have high correlation with MVD.It can reflect the microvessel structure change of the liver cancer tissue and help to judge the curative effect and prognosis of HIFU for liver cancer.

Objective To prepare hepatocyte growth factor(HGF) recombinant protein and confirm its activity preliminarily according to building HGF gene prokaryotic expression vector and transforming into E.coli.Methods Clone HGF inserted into the vector pET-26b(+) to construct prokaryotic expression vector pET-26b(+)-HGF and transform into E.coli Rosseta(DE3).The transformed bacteria induced by IPTG was purified through Ni-NTA resin affinity chromatography frozen-drying after renaturation.Results HGF gene recombinant prokaryotic expression vector pET-26b(+)-HGF was constructed successfully.E.coli Rosseta(DE3) which was transformed into pET-26b(+)-HGF expresses the target protein as the form of inclusion bodies,accounting for 38％ of the total bacterial proteins,and confirmed by Western blot.HGF protein which was purified by Ni-NTA resin affinity chromatography,has a purity of about 95％,and can promote proliferation,migration,and inhibition of apoptosis for human non-small cell lung cancer cell line A549 cells after interaction.Conclusion HGF gene recombinant prokaryotic expression vector pET-26b (+)-HGF was constructed and expressed in transformed E.coli Rosseta(DE3) successfully.They resumed their recombinant HGF protein structure after purification and renaturation,and had biological activity confirmed by in vitro studies.

Background Corneal epithelial abrasion results in corneal ulcer and stroma cloudy evenb irreversible visual impairment.Previous drugs for corneal epithelial injury can only alleviate the inflammatory irritation.So it is very important to seek a drug which regulate the growth of corneal epithelium.Objective This study was to investigate the effects of recombinant human BIGH3 protein eye drops on corneal epithelial abrasion.Methods Fifty right eyes of 50 clean adult New Zealand white rabbits were collected.Two rabbits were sacrificed right away following establishment of corneal epithelial abrasion models (0 hour group).The other 48 rabbits were randomly divided into recombinant human epidermal growth factor (EGF) derivative group (positive control group),normal saline solution group (negative control group),0.25％ or 0.5％ recombinant human BIGH3 protein eye drops group.Corneal abrasion models were created with alcohol corrosion method with a defect area of 7 mm2.The corresponding eye drops were used separately in 4 groups for four times per day after operation.Experimental eyes were examined by the slit lamp microscope,and fluorescein vital staining were performed 12,24,36,48,72 hours after operation.Planimetry was performed and the corneal photographs were analyzed with computer software.The rabbits were sacrificed 12,24,36,48 and 72 hours after operation,respectively,and the histopathological examination of corneal tissue was carried out.Results No obvious irritation response was seen after administered of eye drops in the recombinant human EGF derivative group,normal saline solution group,0.25％ and 0.5％ recombinant human BIGH3 protein eye drops groups.Histopathological examination revealed a full-thickness defect of corneal epithelium after modeling.The defect area was gradually smaller with time lapse,and corneal epithelium migrated from periphery toward the center zone.Corneal epithelial cells increased with time lapse.Compared with normal saline solution group,the defect area of corneal epithelium lessened 12,24,36,48 hours after operation in the 0.25％,0.5％ recombinant human BIGH3 protein eye drops groups and recombinant human EGF derivative group (all at P =0.000),but at 12and 24,36 hours after operation,no significant differences were found between the recombinant human EGF derivative group and normal saline solution group (P =0.321,0.057,0.126).The defect area was smaller in the 0.5％recombinant human BIGH3 protein eye drops group than that of the recombinant human EGF derivative group at various time points (P=0.042,0.039,0.025,0.008).However,significant smaller defect area was exhibited only at 12 hours and 24 hours after operation in the 0.25％ recombinant human BIGH3 protein eye drops group (P=0.047,0.042).No significant differences were seen in corneal defect area at various time points between 0.25％ and 0.5％recombinant human BIGH3 protein eye drops groups (P =0.358,0.259,0.108,0.062).In addition,the corneal defect area was (0.51 ±0.42)mm2 72 hours after operation in the normal saline group;while that in the recombinant human EGF derivative group and recombinant human BIGH3 protein eye drops groups was disappeared.The repairing curves in the recombinant human BIGH3 protein eye drops groups were superior to those of the recombinant human EGF derivative group and normal saline solution group.Conclusions 0.25％ and 0.5％ recombinant human BIGH3 protein eye drops have facilitation effect on the growth of corneal epithelial cells and the healing of corneal injury.

AIM: To test the ability of multifocal visual evoked potential (mfVEP) in the detecting of glaucoma by comparing the mfVEP recorded from normal subjects and glaucoma patients.METHODS: The mfVEP of 32 normal eyes (n =21) and of 58 eyes (n =37) with primary glaucoma were recorded with the Vision Monitor electrophysical apparatus by the second kernel analysis and to determine the correlation of the topographic location between them.RESULTS: There were significant variability (the coefficient of variation was 43.05%) in mfVEP RMS amplitude in the normal subjects; The RMS amplitude of eyes with glaucoma were smaller than that of the normal eyes and significantly statistical difference were found in the relatively center (namely the 0° -10° ring zone) and in superior nasal quadrant (P＜0.05) while there were no significantly statistical differences of the latency time between them.CONCLUSION: The normal subjects have large individual variability of mfVEP responses. The RMS amplitude of the mfVEP of glaucomatous eyes descends, especially in center zone and superior nasal quadrant.

Adult ; Female ; Hand ; surgery ; Humans ; Middle Aged ; Reconstructive Surgical Procedures ; methods ; Toes ; transplantation

Objective To learn the population and infestation rates of cockroaches from 2017 to 2019 in Jiading District of Shanghai, to evaluate the effect of cockroach termination in household, and to provide information for cockroach control. Methods Cockroaches were controlled by dinotefuran baits and clean-up in households.Sticky trap and visual method were employed for density monitoring in farmers markets, supermarkets, hotels, restaurants, hospitals, and residential areas.Visual method was used in households before and after using the insecticide. Results Sticky trap result showed the room infestation rate was 3.24%, mean adhesion rate was 3.29%, the density was 0.06 per board, and the density peak appeared in May.Rate of invasion and density decreased year by year.Blattella germanica was the dominant species, counting for 71.88%.The density, and rate of infestation, as determined by sticky trap method, were the highest in the farmers markets, followed by hospitals and residential areas.Determined by visual method, room infestation rate was 1.16%, and the infestation rate was 4.44%.The peak appeared in January.Infestation rate of the farmers markets was the highest, followed by hospitals and residential areas.By visual method, the room infestation rate was 59.01%, and 48.45% for nymphs.The room infestation and ootheca rates were 54.04% and 17.39%.The rate decreased more than 80% in 30 days after use of the insecticide. Conclusion Infestation rate of cockroach remains at low level in Jiading District.The effect of bait combined with environmental cleaning is remarkable.Future work should strengthen monitoring and control in farmers markets, hospitals and residential areas.

OBJECTIVETo investigate the expression and significance of Cyclin D1 in oral squamous cell ma (OSCC).

METHODSA immunohistochemistry method, Envosion, was employed to test the manifesting Cyclin D1 in pathological slices of 50 OSCC cases and 10 normal cases, and the results was treated with statistical lysis.

RESULTSIn 50 OSCC cases, Cyclin D1 mainly manifested in karyon, and a little in cytoplasm. manifesting rates of Cyclin D1 in the samples was 80.0%, which was significantly higher than the manifesting of 20.0% in normal oral mucous membrane (P < 0.01). The manifestation of Cyclin D1 was correlated with rent pathological grades, clinical phases and lymph node metastasis (P < 0.05).

CONCLUSIONThe abnormal tation of Cyclin D1 is closely related with the occurrence and development of OSCC. Therefore, it can subsidiary index for OSCC treatment and prognosis.

Carcinoma, Squamous Cell ; Cyclin D1 ; Female ; Humans ; Immunohistochemistry ; Lymphatic Metastasis ; Mouth Mucosa ; Mouth Neoplasms ; Prognosis

UNLABELLEDTo identify differentially expressed genes between fetal mesenchymal stem cell (MSC) and adult MSC, especially specified genes expressed in fetal MSC, a cDNA subtractive library of fetal MSC was constructed using suppression subtractive hybridization (SSH) technique. At first, total RNA was isolated from fetal and adult MSC. Using SMART PCR synthesis method, single-strand and double-strand cDNAs were synthesized. After Rsa I digestion, fetal MSC cDNAs were divided into two groups and ligated to adaptor 1 and adaptor 2 respectively. Results showed that the amplified library contains 890 clones. Analysis of 890 clones with PCR demonstrated that 768 clones were positive. The positive rate is 86.3%. The size of inserted fragments in these positive clones was between 0.2 - 1 kb, with an average of 400 - 600 bp.

CONCLUSIONSSH is a convenient and effective method for screening differentially expressed genes. The constructed cDNA subtractive library of fetal MSC cDNA lays solid foundation for screening and cloning new and specific function related genes of fetal MSC.

Cloning, Molecular ; DNA, Complementary ; genetics ; metabolism ; Deoxyribonucleases, Type II Site-Specific ; metabolism ; Fetus ; Gene Library ; Humans ; Mesoderm ; cytology ; metabolism ; Polymerase Chain Reaction ; Stem Cells ; cytology ; metabolism

BACKGROUND AND OBJECTIVEWe previously reported that C-KIT overexpression and mutation exist in biopsy samples of nasopharyngeal carcinoma (NPC). Yet whether Imatinib had an inhibitory effect on the proliferation of NPC in vitro was still unknown. So, this study examined whether sensitivities to Imatinib of other cell lines are different and whether C-KIT expression and mutations exist, to analyze the correlations between them.

METHODSThe expression of C-KIT in NPC cell lines, including CNE-1, CNE-2, Hone-1, C-666, SUNE-1, 5-8F, and nasopharyngeal epithelial (NPE) cell line NP-69, were detected by Western blot. Direct sequencing of polymerase chain reaction (PCR) products was performed to analyze the sequences of C-KIT from the above-mentioned cell lines. Inhibitory effects on proliferation by Imatinib on these cell lines were determined by CCK-8 assay. Pearson product moment correlation and t test were used to analyze the correlation betweeen C-KIT overexpression, C-KIT gene mutation, and the inhibitory effect of Imatinib.

RESULTSCompared with NPE cell line NP-69, NPC cell lines CNE-1, CNE-2, Hone-1, C-666, SUNE-1, and 5-8F had significantly higher levels of C-KIT expression. Heterozygous IVS17+78T>C were found in CNE-1, CNE-2, Hone-1, and NP-69 cell lines, homozygous IVS17+78T>C was found in C-666, and no mutation was found in SUNE-1 or 5-8F. Imatinib had a dose-dependent inhibitory effect on proliferation for CNE-1, CNE-2, Hone-1, C-666, SUNE-1, and 5-8F. No significant correlation between the inhibitory effects of Imatinib, C-KIT overexpression, or C-KIT mutation was found.

CONCLUSIONC-KIT overexpression and intron mutation were found in NPC cell lines and Imatinib had a dose-dependent inhibitory effect on proliferation for NPC cell lines, yet no significant correlation between C-KIT overexpression, C-KIT mutation, or the inhibitory effect of Imatinib was found.

Antineoplastic Agents ; pharmacology ; Benzamides ; Carcinoma, Squamous Cell ; genetics ; metabolism ; pathology ; virology ; Cell Line ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Dose-Response Relationship, Drug ; Epithelial Cells ; cytology ; metabolism ; Herpesvirus 4, Human ; isolation & purification ; Heterozygote ; Homozygote ; Humans ; Imatinib Mesylate ; Introns ; Mutation ; Nasopharyngeal Neoplasms ; genetics ; metabolism ; pathology ; virology ; Nasopharynx ; cytology ; Piperazines ; pharmacology ; Proto-Oncogene Proteins c-kit ; genetics ; metabolism ; Pyrimidines ; pharmacology