Objective To study the methods and efficacy of selective or superselective renal artery embolization for renal neoplasms,abcesses and bleeding. Methods Sixty cases of renal carcinomas and 4 cases of abcesses were all treated with selective renal artery embolization before surgery,and 3 cases of advanced renal carcinomas were treated with selective renal artery embolization alone.Embolization materials for carcinoma and abcesses were mainly gelfoam.Nine cases of renal angiomyolipomas,23 of traumatic renal hemorrhages,1 undergoing extracorporeal shock-wave lithotripsy (ESWL) and 2 nephrolithotomy,all of whom had severe hematuria,were treated with selective or superselective renal segment artery embolization.Embolization materials for hemorrhages were mainly self-blood clots and(or) gelfoam,and the materials for angiomyolipoma were pure alcohol and(or) gelfoam. Results The sizes of carcinomas and abcesses were decreased in all of the 60 cases of renal carcinomas and 4 cases of renal abcesses.The edema around the carcinomas and abcesses became obvious,and bleeding was reduced,which were convenient for operation and prolonged survivals.All the renal bleeding were cured with selective or superselective renal artery embolization except for one traumatic case treated by operation for continuous bleeding.All these cases were followed up for 1 month to 18 years.One case had mild atrophy of kidney and in one case IVU didn't develop.All the others were in good condition. Conclusions Selective or superselective renal artery embolization is a safe,effective approach with less complications and most probability to preserve the diseased kidney. Renal artery embolization as a valuable treatment should be studied and widely applied.

Objective To knock out Asxl2 gene in murine embryonic fibroblast cell line NIH3T3 using CRISPR/Cas9n system. Methods A pair of sgRNAs which targeted exon 5 of Asxl2 gene were designed and subcloned into the pX462 vec?tor. The recombined plasmids were verified by sequencing and transfected into NIH3T3 cell line. Single cells were isolated through serial dilutions, followed by an expansion period to obtain new monoclonal cell lines. The genomic DNA of the new monoclonal cell lines was extracted and a DNA fragment flanked the target site was amplified by genotyping PCR then se?quenced. Lastly, western blotting were applied to confirm whether Asxl2 was successfully knocked out. Results The CRIS?PR/Cas9n plasmids that targeted Asxl2 were successfully constructed. NIH3T3 cells were co-transfected with the two recom?binant constructs. After puromycin selection, subclonal cell lines were obtained and one of them was validated by genotyping PCR-sequencing. Western blotting also confirmed that Asxl2 was completely depleted in the NIH3T3 cell line. Conclu?sion CRISPR/Cas9n plasmids that targeted Asxl2 were successfully constructed therefore a Asxl2 knockout NIH3T3 stable cell line was established via this system.

ObjectiveToinvestigatethekidneyyangdeficiencyconstitutionandhypothalamus-pituitary-adrenal ( HPA) axis dysfunction in Lewis rats .Method Two kidney yang deficiency models were established by subcutaneous injection of hydrocortisone and by adrenalectomy to induce hypothalamic -pituitary-adrenal ( HPA) axis dysfunction in Lewis rats.Wistar rats were used as control and compared with the two types of Lewis rat models of kidney yang deficiency .The degree of kidney yang deficiency of the Lewis rats was evaluated by examination and detection of the general signs , behavior , and neuroendocrine function , and so on .Result Compared with the normal Wistar rats , the body weight of Lewis rats was significantly higher than that of Wistar rats at the same age .The body temperature , urine volume and grip strength were significantly lower than those of Wistar rats ( P<0.01 for all ) .The memory ability of the Lewis rats was slightly decreased.The liver, kidney and adrenal indexes were significantly decreased in the Lewis rats (P<0.01).The levels of serum ACTH, CRH, cGMP, Cort and urine 17OHCS were significantly decreased (P<0.01).Conclusion A very slight deficiency of adrenal cortex function of Lewis rats is caused by genetic inheritance , and Lewis rats have the constitution of congenital kidney yang deficiency ( criticality or prophase of kidney yang deficiency ) .

Objective To construct recombinant adenovirus vector containing Rattus norvegicus microtubule-associated protein 4 gene,and transfect it into the rat cardiac myocytes cultured in vitro.Methods A pair of primers were designed,and full length MAP4 DNA was cloned from rat total mRNA by PCR.The PCR product was double-digested with restriction endonucleases NheⅠ and NocⅠ,and inserted orientationally into pShuttle2.The plasmid of pShuttle2-MAP4 was double-digested with restriction endonucleases NheⅠ and NocⅠ,and inserted BD Adeno-X~(TM) Virul DNA,named pAd2-MAP4.The non-recombinant adenovirus was screened out with PacⅠ, pAd2-MAP4 was linerized with SwaⅠ,and the recombinant virus genome was transfected into HEK293 cell line for packaging and amplification of Ad-MAP4 virus.The recombinant adenovirus was transfected into rat cardiac myocytes and MAP4 was identified by immunohistochemistry.Results The recombinant adenovirus-MAP4 was constructed successfully and the titer was about 2.3?10~(8) pfu/ml.The expression of MAP4 was enhanced at 48 h after the transfection.Conclusion We have successfully constructed a recombinant adenovirus Ad-MAP4 that has enforced the expression of MAP4 in vivo.

OBJECTIVETo explore the proliferation and invasive effects of inhibitors of kinase 4(INK4)(P15(ink4b) and P16(ink4a)/CDKN2) gene protein activation on RKO human colorectal cell in vivo and in vitro.

METHODSRKO human colorectal cell line was exposed to the specific DNA methyltransferase inhibitor 5-Aza-CdR and INK4(P15(ink4b) and P16(ink4a)/CDKN2) protein expression was detected by Western blotting. Soft agar cloning experiment and Transwell chamber assay were used to detect the proliferative and invasive ability in vitro. Tumorigenicity in nude mice was analyzed in vivo.

RESULTSINK4(P15(ink4b) and P16(ink4a)/CDKN2) protein expression of RKO human colorectal cells after exposure to 1×10(-7), 5×10(-7) and 1×10(-6) mol/L 5-Aza-CdR concentrations(A, B, C groups) were 1.13, 1.38, 1.92 folds and 1.11, 1.45, 2.14 folds compared to positive control group respectively. Soft agar cloning experiment showed the number of cell colony significantly decreased from 36.8±5.1(positive control group) to 32.4±7.2, 21.3±5.4 and 19.5±6.4 (3 experiment groups, all P<0.05) respectively. Transwell chamber assay showed that migrated cell number in positive control group(67.4±7.2) was significantly higher than those of 3 experimental groups(35.3±4.6, 29.5±7.3 and 25.3±6.2, respectively). The tumor volume of metastasis model in nude mice was inhibited in experimental groups, but not significantly lower compared to control group (P>0.05). There were significant differences of tumor weight and inhibition rate between control group and 3 experimental groups in nude mice respectively(all P<0.01).

CONCLUSIONINK4(P15(ink4b) and P16(ink4a)/CDKN2) protein activation can inhibit tumor proliferation, migration and suppress the tumor formation ability.

Animals ; Cell Line, Tumor ; Cell Proliferation ; Colorectal Neoplasms ; metabolism ; pathology ; Cyclin-Dependent Kinase Inhibitor p15 ; genetics ; metabolism ; Cyclin-Dependent Kinase Inhibitor p16 ; genetics ; metabolism ; Humans ; Mice ; Mice, Inbred BALB C ; Neoplasm Metastasis ; Neoplasm Transplantation ; Transcriptional Activation

Objective: This study aims to investigate the epidemiological distribution of HFMD and quantify the association of temperature with the incidence of children’s HFMD in Nanjing, China. Methods: Daily counts of HFMD in children under 5 years and daily meteorological variables during 2011-2016 were obtained. Descriptive statistics were used to describe the epidemiological characteristics and distributed lag non linear model (DLNM) was used to assess the associations of temperature on HFMD cases. Results: A total of 104 977 HFMD cases aged 0-5 years were reported in Nanjing during the study period and the male to female sex ratio was 1.49∶1. The average annual incidence was 213.5 per 100 000. A bimodal seasonal pattern was observed and the south and west were found to be the high incidence areas in the city. Of these laboratory confirmed enteroviruses positive cases, 32.5% cases were positive for EV-A71 infections, 29.1% cases were positive for CV-A16 infections and 38.4% cases were positive of other enteroviruses infections. The temperature HFMD relationships were non linear and showed obvious lag effects. The cumulative relative risk presented as an approximately inverted U shape over 14 days and peaked at 25.7 ℃ with value of 2.71(95%CI=1.93-3.81). Subgroup analyses revealed that males and children aged <1 year were more vulnerable to temperature variations. Conclusion Epidemiological characteristics of HFMD among children aged 0-5 years old in Nanjing presented temporal and regional distribution. The temperature has significant impact on children’s HFMD occurrence.

As an effective vehicle for bio-research and for gene therapy, Lentiviral Vector (LV) has been drawn large attention in recent years. However, transcriptional read-through limits its application. In order to understand the extend of LV read-through in chromosome, a reliable method to assess transcriptional read-through rate is needed. Here, we report the method as follows: 293T cells were transfected with the lentiviral transfer vectors which borne with two LTRs at its two ends in order to mimic the state of "proviral vectors" in chromosome. Using the primers specific for 3'U5 and 3'U3, read-through and total transcripts were reverse transcribed, respectively. These two cDNAs were quantified by realtime PCR using the primers and probe specific for 5'end of 3'U3. Read-through rate was then calculated by the division of the two. Meanwhile, read-through product of green fluorescence protein was also analyzed by Fluorescence Activated Cell Sorter. They both reciprocally proved the principal and confirmed that self-inactivated LV appeared higher read-through rate than the wild type one. The method described in this article, therefore, provides a useful technique to study how to reduce read-through rate, and improve the bio-safety of LV.
Flow Cytometry ; Genetic Therapy ; Genetic Vectors ; Green Fluorescent Proteins ; biosynthesis ; HEK293 Cells ; Humans ; Lentivirus ; Transfection

Objective To analyze the influence of different materials on thermal stress field of the bipolar high-frequency electric knife. Methods The electric-thermal coupling simulation analysis was performed for thermal stress field in working process of the bipolar high-frequency electric knife. The influence patterns for 4 kinds of insulating layer materials 304 stainless steel, 316 stainless steel, 317 stainless steel, Ti6Al4V and 4 kinds of electrode materials polypropylene (PP), polyamide-6 (PA6), polycarbonate (PC) and acrylonitrile butadiene styrene (ABS) on thermal stress field and thermal deformation field of insulating layer, electrode and coating of bipolar high-frequency electric knife and myocardial tissues were studied. Results The thermal deformation of myocardial tissues was much higher than that of the components of high-frequency electric knife in period of 2 s for continuous power. In terms of myocardial tissue protection, 304 stainless steel was the best electrode material for high-frequency electric knife. For choosing PP as the insulating layer material, thermal deformation of myocardial tissues caused by thermal stress was the smallest, which could better maintain the integrity of myocardial tissues. Conclusions The manufacturing materials of high-frequency electric knife have an important influence on thermal stress field in its working process. The regularity analysis results can provide guidance for the development of high-frequency electric knife.