In order to provide highly purified preparation of human ieukoregulin (LR) for the exploration of its anticancer mschatiism, crude preparations of LR and lymphotoxin(LT) derived from human spleen cell cultures induced with phytohem-agglutinin and diterp;ne ester l2-O-tetradecanoylphorbol-i3-acetate were purified by using four purification nmhois : (a) Con A-Sepharose 48 affinity chromatography, by which crude human LR or LT was purified to maximum specific activity of 85 333 U/mg protein or 13333 U/mg protein with a recovery of 28.1% or 31.9%, respectively 5 (b) DEAE-Sjphadex A-50 ion-exchange chromatography, by which partially purified LR or LT obtained after Con A-Sepharose 4B affinity chromatography was further purified 2.44-fold or 3.8o-fold with a recovery of 96% or 92%, respectively ; (c) DEAE-22-celluiose ion-exchange chromatography, by which crude human LR or LT was purified to maximum specific activity of 11 294 U/mg protein or 13 176 U/mg protein with a recovery of 87.5% or 89.6%.respectively ; (d) Blue-Sepharose CL-6B affinity chromatography, by which crude human LR or LT was purified to maximum specific activity of 51.89 U/mg protein or 2594 U/mg protein with a recovery of 53.75% or 5t3.G7%, respectively.Our data indicate that Con A-Sepharose 4B affinity chromatography, DEAE-Sephadex A-50 ion-exchange chromatography.DEAE-22-cellulose ion-exchange chromatography and Blue-Sepharose CL-6B affinity chromatography may be useful in the purification of LR or LT.

The cellular sources of leukoregulin (LR) and lymphotoxin (LT) from human peripheral blood leukocytes or spleen cells were examined The ratios of LT activity to LR activity in lymphokine preparations from different individuals were not constant but varied over 640-fold, suggesting that LR and LT activities were mediated by distinctly different entities.Purified LR was used to examine the relative susceptibilities of human or mice cells to LR.Most human tumor cells were very sensitive to LR while human normal cells and mice normal or tumor cells were less sensitive or resistant.Crude preparations of human interferon (Hu IFN)-? or Hu IFN-? were more cytotoxic to human tumor cells than partially purified natural Hu IFN-? or highly purified rHu IFN-?D, and crude or purified LR were more cytotoxic to human tumor cells than Hu IFN-? Hu IFN-?.Hu IFN-? activity but not cytotoxic activity could be removed from preparatons containing Hu IFN-?,LR and LT activities with the aid of monoclonal antibody to Hu IFN-?. Natural killer cytotoxic factor (NKCF) supernatants from the co-culture of human spleen cells and NK susceptible cell line K562 or Molt-4 were strongly cytotoxic to various tumor cell lines.The NKCF activity was completely adsorbed after incubation with Molt-4 cells and completely inactivated by incubating at pH2 for 24 h.The putative receptor for LR or LT was studied using adsorption assay.The present study reveals that human LR activity can be adsorbed from crude LR supernatants by incubation with native or formalin-treated K562 or SMMC-7721 cells at 37℃ or 4℃; LT activity can be adsorbed with L929 cells.Therefore, LR appears to be a cytotoxic/cytostatic factor that is distinct from LT, 1FN and NKCF.

A simple, rapid and accurate method is described for the determination of icariin in Fuchun Tablets by reversed-phase high-efficiency liquid chromatography. The samples were first extracted with 70% alcohol for 50 minutes The resulting solutions were chromatographed on aYWG-C18 column.By using a mobile phase of methanoltetrahydrofuran-water (26.5:15.9:56.7) and a detection wavelength at 274 nm, the recovery for 101.02%, CV=2.4%,are obtained.The method can be employed for the quality control of Fuchun Tablets.

Object To optimize the best technical parameters through controlling the different factors. Methods Taking spherical degree and pill weight as index, the above factors were observed by orthogonal test. Results PEG4000 as matrix, methyl silicon oil as refrigerant. Internal and external diameter of burette are 4 1 mm and 6 1 mm, dropping into the refrigerant that composed of 40 ℃—50 ℃,10 ℃—30 ℃,0 ℃—4 ℃ by 20 ℃—30 ℃ droppings per minute. Conclusion The highest finished product with good quality can be got through this process.

Objective: To observe the effect of electroacupuncture (EA) at Lower He-Sea points on the expression levels of interleukin-1β (IL-1β) in the serum and gallbladder tissues, and nuclear factor-κB (NF-κB) in gallbladder tissues of the guinea pigs with acute cholecystitis (AC), and to explore whether Yanglingquan (GB 34), the Lower He-Sea point pertaining to Dan Fu (gallbladder), is relatively specific for the Dan Fu (gallbladder) disorders. Methods: Eighty-two healthy guinea pigs were randomly divided into 6 groups according to the random number table method, a blank group, a model group, a Yanglingquan (GB 34) group, a Zusanli (ST 36) group, a Shangjuxu (ST 37) group, and a Xiajuxu (ST 39) group, with 12 guinea pigs in the blank group while 14 in the other groups, respectively, half males and half females in each group. Except for the blank group, guinea pigs in the other groups were injected with E. coli into the gallbladder to establish AC models. Guinea pigs in the blank group were fed routinely without special treatment; those in the model group were daily tied up for 30 min without EA treatment; those in the 4 groups receiving EA treatment were acupunctured at the corresponding Lower He-Sea points after daily binding and stimulated with the SDZ-V EA instrument. After successful modeling and treatment for 5 d, blood was collected from the abdominal aorta of the guinea pigs, and the gallbladder tissues in each group were isolated for hematoxylin-eosin (HE) staining to observe the morphological changes. Enzyme-linked immunosorbent assay (ELISA) was used to detect the serum IL-1β level, and immunohistochemistry (IHC) was used to detect the expression levels of NF-κB and IL-1β in gallbladder. Results: On the 3rd day after modeling, the guinea pigs in the five groups with modeling were mentally depressed with decreased appetite, significantly reduced activities, slouch, lassitude, slack and matted fur, and loose stools; two guinea pigs were selected from each group (one male and one female, not included in the final statistics) to isolate the gallbladder after sacrifice; macroscopic observation showed that the gallbladder wall was differently thickened; the bile color was dark green and opaque with particles suspended or accumulated; light microscope observation showed that the submucosal blood vessels of the gallbladder were congested, along with mucosal edema, ulceration, necrosis, shedding, and a large number of inflammatory cells infiltrating in the lamina propria, indicating that the AC model was successfully prepared. Compared with the model group, the gallbladder tissue injuries of the four groups receiving EA treatment were all differently repaired, the serum IL-1β levels were reduced (P<0.01 or P<0.05), and the IL-1β levels in the gallbladder tissues were reduced (P<0.01 or P<0.05). Compared with the model group, the NF-κB expression level in the Yanglingquan (GB 34) group was significantly reduced (P<0.01), but was not statistical different in the Zusanli (ST 36) group, Shangjuxu (ST 37) group and Xiajuxu (ST 39) group (all P>0.05). Compared with the Yanglingquan (GB 34) group, the gallbladder tissues of the Zusanli (ST 36) group, Shangjuxu (ST 37) group and Xiajuxu (ST 39) group were more severely damaged, and the expression levels of serum IL-1β, the NF-κB and IL-1β in the gallbladder tissues were increased (P<0.01 or P<0.05). Intervention effect of Yanglingquan (GB 34) on AC guinea pigs was superior to that of Zusanli (ST 36), Shangjuxu (ST 37) and Xiajuxu (ST 39). Conclusion: EA at the Lower He-Sea points of the stomach, large intestine, small intestine and gallbladder can produce curative effects on AC guinea pigs and reduced the inflammatory symptoms. Intervention effect of Yanglingquan (GB 34) on AC guinea pigs is superior to that of Zusanli (ST 36), Shangjuxu (ST 37) and Xiajuxu (ST 39). The mechanism of EA at Yanglingquan (GB 34) in treating AC may be regulating IL-1β and NF-κB to control the inflammatory response and improve the gallbladder tissue damage.

Humans ; Poisoning ; prevention & control ; therapy ; Solvents ; poisoning

Humans ; Mastoiditis ; diagnosis ; surgery ; Otitis Media ; diagnosis ; surgery

Objective:To investigate the expressions and their clinical significance of vascular endothelial growth factor (VEGF) and extracellular matrix (ECM) components in non-small cell lung cancer (NSCLC). Methods:Expressions of VEGF and ECM components (fibronectin, FN and collagen Ⅳ, cⅣ) in 50 cases of NSCLC tissues and 20 cases of normal lung tissues were detected by immunohistological analysis. Their relationship with clinical features of NSCLC and the correlation of expression of VEGF and Fn and cⅣ were analyzed.Results:The positive expression rates of VEGF, Fn, and cⅣ were 96%, 78%, and 50% in NSCLC tissues. The expressions of VEGF and Fn were significantly higher than those in normal lung tissues (P<0.05). The expression of Fn and over-expression of VEGF were associated with lymph node metastasis (r=1.00, P<0.001). The survival rate of patients with over-expression of VEGF was greatly lower than that with weak expression of VEGF (P=0.022). The survival rate of Fn-negative patients was markedly higher than that of Fn-positive patients (P=0.046). Conclusion:VEGF and ECM component Fn were highly expressed in NSCLC, which correlated with lymph node metastasis and survival rate. Expression of ECM and VEGF had positive correlations, suggesting that ECM might be one of the anti-angiogenesis targets for tumor therapy.

Objective:To make a prospective study on the effectiveness and safety of toremifene (TOR) combined with novelbine/cisplatin (NP) in the treatment of patients with advanced non-small cell lung cancer (NSCLC) whose first line platinum-based chemotherapy was failure. Methods:Forty-four patients with stage ⅡB-Ⅳ NSCLC, who failed in the first line cisplatin-based chemotherapy from January 2004 to February 2006, were enrolled in this study. All the patients received TOR combined with NP second line chemotherapy for two cycles. The response rate and adverse reaction were evaluated. The survival rate was analyzed.Results:The 44 patients received average 1.8 cycles of chemotherapy (1-3 cycles). The response of 37 patients could be evaluated including 21 patients who received NP regimen before and 16 patients who received platinum-based chemotherapy. After second line therapy, 4 of the 37 patients had partial response (PR), 19 had stable disease (SD), 14 had progressive disease (PD), and no patient had complete response (CR). The total response rate (CR+PR) was 10.8% (4/37). The disease-controlling rate (CR+PR+SD) was 62.2% (23/37). The response rate and disease-controlling rate of squamous cell lung cancer (SCC) were 27.3% (3/12) and 72.7% (8/12), which were significantly higher than adenocarcinoma [0% (0/18) and 44.4% (8/18), P<0.05]. The median survival time was 8.2 months, the median time for SD was 4.0 months (1.0-10.2 months), and the 1-year survival rate was 24.4%. The median survival time and 1-year survival rate of SCC patients had no significant difference compared with adenocarcinoma patients (9.2 vs 7.1 months; 33.3% vs 27.7%, P＝0.72). There was no significant difference in survival rate between male and female patients. One patient stopped therapy for liver function injury (hyperbilirubinemia). The adverse reactions induced by chemotherapy mainly included gastrointestinal reaction, bone marrow suppression, and liver function injury. No serious adverse reaction occurred. Conclusion:The clinical efficacy of second line TOR combined with NP regimen is similar with the first line regimen for NSCLC patients, especially for SCC patients. The frequency of adverse reaction is not increased.

Objective:To investigate the molecular mechanism underlying apoptosis of lung adenocarcinoma cell line A549 induced by rapamycin, a type of mammalian target of rapamycin (mTOR) signal pathway inhibitor, combined with hypoxia treatment. Methods:Rapamycin was applied to treat A549 cell line under hypoxia for 16 h. Then the discrepancy of cell apoptosis between treated and untreated control was compared by Hochest 33258 staining. After being treated with rapamycin combined with hypoxia, the mRNA and protein expressions of survivin, a kind of anti-apoptotic molecule, were tested by real-time PCR and Western blotting. Western blotting was employed to test the difference of hypoxia-inducible factor (HIF)-1α expression level before and after rapamycin was added. In additon, chromatin immunoprecipitation(ChIP) assay was further applied to explore the mechanism underlying the effects of rapamycin under hypoxia in inducing apoptosis of A549 cells. Results:Rapamycin induced apoptosis of A549 cell line under hypxia in vitro. In A549 cell line, the expressions of survivin at mRNA (P<0.01) and protein levels, were both decreased by rapamycin combined with hypoxia treatment. Besides, ChIP assay revealed that rapamycin inhibited the expression of survivin at the transcriptional level through decreasing the hypoxia-induced up-regulation of HIF-1α and its binding with the promoter of survivin gene. Conclusion:Rapamycin exerted anti-tumor effects by inhibiting the expression of HIF-1α and decreasing the binding of HIF-1α with the promoter of survivin gene, thereby down-regulating the increase in the expression of apoptosis inhibiting gene survivin.