Comparison studies of recombinant prourokinase(pro-UK) in various host cells,and expression vectors were carried out. Expression levels of vectors constructed in this study in different cell lines were compared. Mammalian cells expressing pro-UK were established. The levels of pro-UK expression in recombinant Namalwa, Vero and Sp2/0 cells are 200,12.5 and 50 IU/( 106 cells 24h). pro-UK purities separated by immunoaffinity chromatograph are above 90 %. Immunoabsorbent assay showed the ratio of pro-UK in CHO cells is lower than that from Vero and Namalwa cells. This study provide new host cells for pro-UK production.

BACKGROUNDThere is no validated blood biomarker available for glioma management. Invasive growth is the key feature of glioma. We assessed the clinical usefulness of plasma tissue inhibitor of metalloproteinase 1 (TIMP-1), which has less molecular weight than metalloproteinases, as a potential blood biomarker for glioma.

METHODSA total of 285 patients and 59 normal subjects were studied. Plasma concentration of TIMP-1 was measured with enzyme-linked immunosorbent assay. Plasma TIMP-1 was compared between normal and glioma patients, between patients with different pathological grades, and between patients with different prognoses. Longitudinal changes in plasma TIMP-1 during treatment were also evaluated. Plasma matrix metalloproteinase (MMP)-9 level was also assayed and its clinical usefulness was compared with that of TIMP-1.

RESULTSPlasma TIMP-1 and MMP-9 were both increased in glioma patients compared with normal controls (TIMP-1: P < 0.001; MMP-9: P = 0.007). Plasma TIMP-1 increases with increased tumor grade. In Grade IV gliomas, plasma TIMP-1 significantly increased after "successful removal" of the tumor (paired samples t-test, before operation vs. during chemotherapy without recurrence, t = -2.131, P = 0.038), but did not change significantly at the time of tumor recurrence (during chemotherapy without recurrence vs. after tumor recurrence, t = -0.652, P = 0.632). High plasma TIMP-1 level correlated with better survival in Grade IV glioma patients (hazard ratio: 0.550, 95% CI: 0.101-1.000, P = 0.036). In Grade IV gliomas, patients with higher plasma TIMP-1 had significantly longer survival time than those with lower plasma TIMP-1 level (25.23 vs. 18.95 months, log-rank P = 0.045). Plasma MMP-9 did not show significant association with either the pathological grade or the prognosis of glioma patients.

CONCLUSIONSPlasma TIMP-1 is associated with the diagnosis and prognosis of glioma patients. It appears to have better usefulness for guiding clinical decision making than plasma MMP-9. Further studies in an expanded patient population are needed to better define its clinical usefulness.

Adult ; Biomarkers, Tumor ; Case-Control Studies ; Female ; Glioma ; blood ; diagnosis ; Humans ; Male ; Middle Aged ; Tissue Inhibitor of Metalloproteinase-1 ; blood

The only difference of primary structure between single-chain prourokinase (pro-UK or scu-PA) and two-chain urokinase (UK or tcu-PA) is the cleavage of a single peptide bond (Lys158-Ile159) and transform scu-PA into its active two-chain form. A 13-peptide (Thr-Leu-Arg-Pro-Arg-Phe-Lys-Ile-Ile-Gly-Gly-Glu-Cys), which spans the cleavage peptide bond, was synthesized and linked to KLH (Keyhole limpet hemocyanin). The Balb/c mice were immunized by the conjugated protein with proper adjuvant. According to the Kohler and Milstein's methods, a hybridoma cell line G7 secreting monoclonal antibody specific for scu-PA was obtained. The anti-scu-PA McAb, purified from the supernatant of porous microcarrier hybridoma cell culture, was conjugated to CNBr-activated Sepharose 4B to prepare an immuno-affinity chromatography column. The u-PA was purified only by this affinity column from the supernatant of cultivating the u-PA-producing recombinant CHO cell, the u-PA recovery ratio is 90.4%, the purification factor was about 50, with the specific activity of 1.2 x 10(5) IU/mg, the scu-PA ratio in the u-PA product was 96.3%. Compared to immuno-affinity chromatography, the 3-step process for purifying u-PA (cation-exchange column, gel filtration column and benzamidine affinity column) has a u-PA recovery ratio of about 65%, with a specific activity of 1.0 x 10(5) IU/mg, and an scu-PA ratio of about 90%. These results showed that immuno-affinity chromatography is simple to recover u-PA and effective to separate scu-PA from tcu-PA.
Animals ; Antibodies, Monoclonal ; immunology ; isolation & purification ; Chromatography, Affinity ; Enzyme-Linked Immunosorbent Assay ; Mice ; Mice, Inbred BALB C ; Recombinant Proteins ; immunology ; isolation & purification ; Urokinase-Type Plasminogen Activator ; immunology ; isolation & purification

OBJECTIVETo investigate the clinical effects of sialoendoscopically-assisted sialolithectomy for the sialolithiasis in the hilum of the submandibular glands.

METHODSBetween December 2005 and March 2011, treated 80 cases of radiography-verified sialiolithiasis in the hilum of the submandibular glands, The patients included 42 males and 38 females aged from 13 to 68 years. All these patients underwent sialoendoscopic observation and sialoendoscopically-assisted sialolithectomy and were followed up periodically for 3 - 6 months after operation. The success rate of stone removal, postoperative complications and clinical effects were analysed.

RESULTSThe stones were completely removed in 71 cases, and almost completely removed in 5 cases, with a successful rate of 95% (76/80). Among 76 successful cases, 8 were treated by basket entrapment, 59 by intraoral open surgery and 9 by both of these two techniques. Within 3 - 6 months' follow-up, 1 case suffered temporary lingual nerve parenthesis and two suffered ranula formation.

CONCLUSIONSSialoendoscopically-assisted sialolithectomy is a safe and effective gland-preservation technique for the patients with the hilum of the submandibular glands.

Adolescent ; Adult ; Aged ; Endoscopy ; methods ; Female ; Follow-Up Studies ; Humans ; Male ; Middle Aged ; Natural Orifice Endoscopic Surgery ; adverse effects ; Ranula ; etiology ; Salivary Duct Calculi ; surgery ; Salivary Ducts ; pathology ; Submandibular Gland ; surgery ; Young Adult

Objective To compare the histologic grade between biopsy and postoperative specimen in bladder urothelial carcinoma,and approach the state and the reasons of underestimate the histologic grade preoperative.Methods We retrospectively 82 cases of urothelial carcinoma at the Third Affiliated Hospi- tal of the Sun Yat-Sen University.For all the cases in this study,the histologic grade,using the 1998 World Health Organization and International Society of Urological Pathologists(WHO/ISUP)classification,was i- dentical when the biopsy specimen and postoperative specimen were compared.Results In this study,35 cases,28 cases and 19 cases were G_1、G_2、G_3 by biopsy preoperative,respectively;while 22 cases,32 cases、28 cases were G_1、G_2、G_3 postoperative,respectively.There were 24 cases(29.3%)underestimate the histo- logic grade by biopsy preoperative in the 82 cases,while 4 cases(4.9%)overestimate preoperative.The state of underestimate the histologic grade is correlated with the location of biopsy,tissue dose and the conser- vation of pathology judgment.Conclusions There were 24 cases(29.3%)underestimate the histologic grade by biopsy preoperative.We should pay more attention to this state of underestimate the histologic grade preoperative in the treatment of bladder urothelial carcinoma.

Objective To elaborate on the classification of chronic obstructive parotitis (COP) based upon endoscopic and sialographic appearances,and to evaluate the results of endoscopic intervention.Methods Forty-three patients with COP underwent endoscopic exploration and intervention procedures.They included 21 patients with bilateral COP and 22 patients with monolateral COP.The images of sialogaphy and endoscopy were analyzed and classified.The treatment options included saline irrigation,mechanical dilatation and balloon dilatation.All patients were followed up periodically after treatment.Results COP can be classified as:① mild lesion:the main duct has stenosis and ectasia,whereas the 1.1 mm endoscope can pass through easily ; ② moderate lesion:the main duct has one point of severe stricture which can not be passed by the 1.1 mm endoscope; ③ severe lesion:the main duct has two points of severe strictures or diffused strictures.The duration of follow-up ranged from 3 to 17 months.One patient with bilateral COP was lost of follow-up.The clinical results included good in 33 glands,fair in 18 glands and poor in 11 glands,with a total effective rate of 82％ (51/62).Conclusions Endoscopic procedure is valuable for the diagnosis and treatment of COP.

Objective To investigate the value of endoscopy in the diagnosis and treatment of sialolithiasis. Methods Diagnostic and interventional sialoendoscopic procedures were Derformed in 52 patients with sialolithiasis(43 submandibular glands and 9 parotid glands).Results Of the 34 sialoliths in the anterior and/or posterior part of the Wharton's duct,24 were removed with basket retrieval:2 removed with open surgery and basket retrieval,and 8 removed with open surgery under the guidance of endoscopy.Eight sialoliths in the hilum of the Wharton'S duct were treated with open surgery.Of the 9 stone cases in the Stensen's duct,3 was removed with basket retrieval,3 was removed after opening-up of the ostium.1 was treated with basket capturing and open surgery.The obstructive symptoms were improved in these cases during 1-24 months' follow-up. Conclusions Sialoendoscopy is a minimal invasive and efficacious technique for the diagnosis and treatment of sialolithiasis.

A novel technique was developed to deal with apoptosis in large-scale animal cell culture. By means of replacing part of Cytopore porous microcarriers at regular intervals, a rCHO cell line, which produces urokinase-type plasminogen activitor(u-PA), was cultivated continuously with serum-free medium in a 30L stirred tank for 91 days. The cell density was maintained at (1.3～2.6)×107/mL, and > 90 % of cells was viable. In order to reduce the effect of cell density on cell growth and expression,a cyclic pressure oscillation was exerted on a 7.5L reactor headspace to enhance cell expression at high cell density to a certain extent. During the 67 days of medium-replacement culture, the maximal cell density reached 2.64×107/mL, and cell viability was always kept above 95 % when combined with microcarrier-replacement. Compare to control culture, culture with cyclic pressure oscillation could enhance cell expression level and reduce the ratio of glucose metabolized anaerobically to produce lactate. With four-step purification process, about 80 g u-PA(～90 % scu-PA)was recovered from～2100 liters supernatant which contained～135 g u-PA.

Lianhua Qingwen preparation （LHQW） is a Chinese traditional patent medicine approved by China Food and Drug Administration （CFDA）， and divided into two dosage forms， namely capsules and granules. Based on TCM theory， its therapeutic functions are contagion-clearing， detoxification， antipyretic， and lung-ventilating regulation， with influenza as its indication. In this paper， its pharmacological activities were reviewed. LHQW had a significant anti-virus effect characterized by a broad-spectrum pattern. It was reported that it not only possessed definitely suppressive effect on a series of influenza viruses， respiratory syncytial virus， coxsackie， enterovirus， herpes simplex virus，but also displayed a significant inhibitory effect on both the new corona pneumonia virus （SARS-CoV-2） and SARS coronavirus （SARS-CoV）. Studies showed that LHQW has obvious anti-inflammatory effects on a variety of inflammation models. It can significantly increase the delayed hypersensitivity of immunocompromised mice （caused by hydrocortisone） against 2， 4-dinitrofluorobenzene， and improve their cellular immune function. It can improve the phagocytosis function of peritoneal macrophages， the serum hemolysin antibody level and the humoral immune function of mice with a low immune function， with a immunomodulatory effect. In addition， LHQW has therapeutic effects on the symptoms induced by respiratory tract infections， such as fever， cough and phlegm， so as to block the vicious circle of multiple pathological links of the disease， and bring the advantages of multi-target， multi-link and multi-approach overall treatment of TCM into play.

OBJECTIVETo investigate the risk factors for the development of congenital anal atresia in neonates.

METHODSA total of 70 neonates who were admitted to 17 hospitals in Foshan, China from January 2011 to December 2014 were enrolled as case group, and another 70 neonates who were hospitalized during the same period and had no anal atresia or other severe deformities were enrolled as control group. Univariate and multivariate logistic regression analyses were used to investigate the risk factors for the development of congenital anal atresia.

RESULTSThe univariate analysis revealed that the age of mothers, presence of oral administration of folic acid, infection during early pregnancy, and polyhydramnios, and sex of neonates showed significant differences between the case and control groups (P<0.05). The multivariate logistic regression analysis revealed that infection during early pregnancy (OR=18.776) and male neonates (OR=9.304) were risk factors for congenital anal atresia, and oral administration of folic acid during early pregnancy was the protective factor (OR=0.086).

CONCLUSIONSInfection during early pregnancy is the risk factor for congenital anal atresia, and male neonates are more likely to develop congenital anal atresia than female neonates. Supplementation of folic acid during early pregnancy can reduce the risk of congenital anal atresia.

Anus, Imperforate ; etiology ; Female ; Humans ; Infant, Newborn ; Logistic Models ; Male ; Pregnancy ; Risk Factors