BACKGROUND/AIMS: To investigate the differential expression of RING finger (RNF) proteins in Barrett esophagus (BE) and esophageal adenocarcinoma (EAC). METHODS: The differential expression of RNFs in normal esophagus (NE), BE, and EAC was screened using microarray assay. Real-time quantitative polymerase chain reaction (PCR), tissue microarray assay, and Western blot analysis were independently performed to detect the mRNA and protein expression of screened RNFs. RESULTS: The expression of nine RNFs in the BE or EAC was 2-fold higher than those in NE. Among these proteins, the RNF32 and RNF121 expression in BE was 20.3-fold and 16.4-fold higher, respectively, than that in NE, and the expression of RNF24, RNF130, RNF141, RNF139, RNF11, RNF14, and RNF159 was upregulated more than 2-fold compared with NE. The expression of nine RNFs was not only upregulated in the EAC but was also positively related to the RNF expression in BE. The PCR results also indicated increased expression of these RNFs in BE and EAC compared to NE. Furthermore, the mRNA expression of all RNFs, except for RNF141 in EAC, was dramatically higher than those in the BE. Similar results were also obtained from the Western blot analysis. CONCLUSIONS: A total of nine RNFs play critical roles in the progression of BE to EAC.
Adenocarcinoma/*enzymology/genetics ; Adult ; Aged ; Barrett Esophagus/*enzymology/genetics ; Carrier Proteins/genetics ; Disease Progression ; Esophageal Neoplasms/*enzymology/genetics ; Female ; Gene Expression Profiling ; Humans ; Intracellular Signaling Peptides and Proteins/genetics ; Male ; Middle Aged ; Proteins/genetics ; *RING Finger Domains ; Receptors, Cell Surface/genetics ; Ubiquitin-Protein Ligases/genetics/*metabolism

OBJECTIVETo provide experimental basis for extending the indications of arsenic trioxide (As(2)O(3)) in clinical application.

METHODSMTT assay was used to detect the cytotoxicity of As(2)O(3) in combination with daunorubicin (DNR), cytosine arabinoside (Ara-C), harringtonine (H) and vincristine (VCR) respectively on leukemic cells from 23 newly diagnosed cases with acute non-promyelocytic leukemia (ANPL) and 16 cases of relapsed, refractory ANPL.

RESULTS(1) As(2)O(3) inhibited the growth of leukemic cells from both newly diagnosed or relapsed and refractory ANPL patients, and there was no statistical difference in cytotoxicity of the patients in the two groups [(12.6 +/-7.7 compared with 10.1 +/-6.2)%, P<0.05]. (2) There was no correlation between the cytotoxicities of As(2)O(3) and Ara-C, H or VCR (P<0.05), but a linear correlation between As(2)O(3)and DNR was found (r=0.432, P<0.05).(3) Additivity and synergism of the cytotoxicity was found in most of the ANPL patients when As(2)O(3) was combined with the four chemotherapy drugs and the combination of As(2)O(3) with DNR or VCR enhanced the cytotoxicity significantly (P<0.05).

CONCLUSIONThe results indicate that As(2)O(3) might be used in treatment of newly diagnosed or relapsed and refractory ANPL patients;and the combination of As(2)O(3) with DNR or VCR may enhance its therapeutic efficacy.

Acute Disease ; Adolescent ; Adult ; Aged ; Antineoplastic Combined Chemotherapy Protocols ; therapeutic use ; Arsenicals ; administration & dosage ; Child ; Drug Resistance, Multiple ; Drug Resistance, Neoplasm ; Female ; Humans ; Leukemia ; drug therapy ; pathology ; Male ; Middle Aged ; Oxides ; administration & dosage

Aim To establish a UPLC method for the determination of the concentration of hainanolidol in plasma of rats, and study the pharmacokinetics of hain-anolidol in rat plasma after single dose i. v. administra-tion of hainanolidol (1, 2, 4 mg·kg-1). Methods The UPLC method for the determination of hainanolidol in rat plasma was established using hainanolide as in-ternal standard. The mobile phase was methanol-water (47 ∶ 53), the flow rate was 0.17 mL·min-1, and the detection wavelength was UV 326 nm. The plasma concentration of hainanolidol in rats was determined by UPLC after single-dose intravenous injection in rats with 1, 2 and 4 mg·kg-1of hainanolidol, and the pharmacokinetic parameters were calculated by DAS2.1. Results The result of calibration curve was linear over the range of 0.05 ~10.00 mg·L-1( r = 0.999 6) . The lower limit of quantification was 0.05 mg·L-1. The intra-day and inter-day precision were both lower than 5% , and the extraction recoveries were higher than 85% , respectively. The validated method was successfully applied to the pharmacokinetic study after i. v administration of hainanolidol in rats with do-ses of 1, 2 and 4 mg·kg-1. The T1/2was (39.82 ± 0.92), (40.11 ± 0.79) and (41.61 ± 2.07) min, respectively. The AUC0-twas ( 65.77 ± 1.08 ) , (130.48 ± 1.26) and (268.75 ± 1.24) min·mg· L-1, respectively. Conclusion A simple and specific UPLC method for the analysis of hainanolidol is suc-cessfully developed, which could be applied to phar-macokinetic study in rat plasma.

Objective： The current study was designed to explore the role of telomerase and its subunits in the process of gastric carcinogenesis. Methods： Telomerase activity was detected with telomeric repeat amplification protocol (TRAP) and its subunits were examined by RT PCR. Results： Telomerase activity was noted positive in 24.6％ (14/57) for chronic atrophic gastritis, 38.9％ (7/18)for intestinal metaplasia, 37.5％ (3/8) for dysplasia and 92.3％ (62/65) for gastric carcinoma, respectively. On the contrary, no telomerase activity was detected in normal mucosa. The frequency of telomerase activity in gastric carcinoma was significantly higher than that in chronic atrophic gastritis, intestinal metaplasia and dysplasia (P< 0.01). Telomerase activity in gastric carcinoma was not related with clinicopathological parameters. Telomerase subunits, TP1 and hTR, were expressed in most gastric mucosas and no significant difference was found among different groups. However, another telomerase subunit,hTRT, was mainly detected in gastric carcinoma and partially premalignant lesions or diseases and the expression of hTRT paralleled to the expression of telomerase. Conclusions： Telomerase activitymay play an important role in the early stage of gastric carcinogenesis. hTRT may be not only a good diagnostic parameter but also a target for gene therapy in gastric cancer because hTRT expression parallels to telomerase activity.

AIMTo investigate the effect of cocaine on apoptosis and caspase-3 activity in germ cells in male rats at different ages.

METHODSCocaine hydrochloride was given (15 mg/kg body weight s.c.) to male Sprague-Dawley rats of 3 weeks (n = 8), 6 weeks (n = 8) and 12 weeks (n = 8) of age, daily for 28 Days. The serum levels of follicle stimulating hormone (FSH), luteinizing hormone (LH), prolactin (PRL), testosterone (T) and estrogen (E2) were assayed, and the DNA fragmentation of germ cells was determined by gel eletronphoresis. The cell cycle, apoptosis and caspase-3 activity of germ cells were tested by flow cytometry.

RESULTSAfter the 28-day cocaine treatment, testes weight of the 3-week-old rats, the testes and body weights of the 6-week-old rats were decreased significantly compared to those of their corresponding controls (P < 0.05). The serum level of T was decreased significantly in the 3-week-old and 6-week-old rats, and the serum level of PRL was also decreased significantly in 12-week-old rats compared to the controls (P < 0.05). In all the three cocaine-treated groups, the isolated DNA displayed a clear ladder pattern, especially in the 6-week old rats. The number of apoptosic germ cells increased significantly in 3- and 6-week-old rats treated with cocaine (P < 0.05). The caspase-3 activity in all three groups increased significantly compared to the controls (P < 0.05), especially in the 6-week-old rats.

CONCLUSIONCocaine exposure for 28 Days leads to significant damage to male gonad and apoptosis elevation in testes of rats of different ages, especially in those of 6 weeks of age. The increase in caspase-3 activity might be a key pathway related to the early stage of apoptosis as the mechanism of cocaine-induced germ cell loss.

Aging ; physiology ; Animals ; Caspase 3 ; Caspases ; drug effects ; metabolism ; Cell Cycle ; drug effects ; Cocaine ; pharmacology ; Estrogens ; blood ; Follicle Stimulating Hormone ; blood ; Luteinizing Hormone ; blood ; Male ; Prolactin ; blood ; Rats ; Rats, Sprague-Dawley ; Spermatozoa ; cytology ; drug effects ; physiology ; Testis ; drug effects ; pathology ; Testosterone ; blood

Purpose To clarify the effect of adenosine on brain metastasis of lung cancer and the possible mechanism of adenosine promoting brain metastasis of lung cancer. Methods Western blot was used to dynamically detect the expression level of hypoxia inducible factor-1 (HIF-1) in lung cancer cells and tight junction protein ZO-1 in brain microvascular endothelial cells on blood-brain barrier. The content of adenosine in lung cancer cell culture was determined by ELISA. Fluorescence analysis was used to detect the changes of permeability of the blood-brain barrier model in vitro. Hemocytometer counts the number of A549 lung cancer cells in Transwell's lower chamber. Results The expression level of HIF-1 in lung cancer cells and the content of adenosine in lung cancer cell culture reached the highest level when lung cancer cells were deprived of oxygen for 12 hours. At the same time, the expression level of ZO-1 protein in the blood-brain barrier was the lowest, the blood-brain barrier permeability was the highest (7.11), and the number of lung cancer cells passing through the blood-brain barrier model was the highest (84.6). The permeability of the blood-brain barrier model increased after the action of adenosine, and its change trend was consistent with the effect of hypoxic lung cancer cell culture solution. Conclusion Hypoxia can induce the lung cancer cell to release adenosine, the increased adenosine can reduce the expression of tight junction protein ZO-1 in blood brain barrier, which leads to the increase of permeability of blood-brain barrier and eventually lead to brain metastasis of lung cancer.

Objective To compare the diagnostic value 18F-fluorothymidine (FLT) and 18F-fluorodeoxyglucose (FDG) PET/CT in detecting lymph node metastases of untreated thoracic esophageal carcinoma. Methods Twenty-two patients with thoracic esophageal squamous cell carcinoma underwent both 18F-FLT and 18F-FDG PET/CT before surgery. The imaging results of the two modalities in detecting regional lymph node metastases were compared prospectively with the pathologic findings. The X2-test was used with SPS S 13.0. Results All patients underwent esophagectomy and lymphadenectomy. The metastatic lymph nodes were found in 16 patients, from which 47 of 424 excised nodes were positive by pathologic examination. False positive results were 14 while false negative 8 on 18F-FDG PET/CT. In contrast, false positive results were only 3 but false negative were 12 on 18 F-FLT PET/CT. The sensitivity, specificity, accuracy,negative predictive value, and positive predictive value were 74.47% ( 35/47 ), 99.20% ( 374/377 ),96.46% (409/424), 96.89% ( 374/386 ) and 92.11% ( 35/38 ) respectively for 18 F-FLT PET/CT, whereas the corresponding values were 82.98% (39/47), 96.29% (363/377), 94.81% (402/424), 97.84%(363/371 ) and 73.58% (39/53) respectively for 18 F-FDG PET/CT (X2 = 0.572, 6.018, 1.017, 0.348,3.852, P＞0. 05, ＜0.05, ＞0.05, ＞0.05 and ＞0.05). Conclusions Compared with 18F-FDG PET/CT, 18F-FLT PET/CT may be less sensitive but more specific for the detection of lymph node metastases of thoracic esophageal carcinoma.

Objective To determine the efficient cut-off points of fasting fingertip blood glucose test for undiagnosed diabetes mellitus(DM), impaired glucose tolerance(IGT), and impaired fasting glucose(IFG)in community-based residents aged above 45 years old. Methods A cluster-randomized study was conducted from May 2008 to January 2009. A total of 3250 subjects aged above 45 years in two communities of Baoding city received questionnaire investigation and tested for fingertip blood glucose. Those subjects whose capillary blood glucose level ≥5.1 mmol/L were subjected to 75 g oral glucose tolerance test. Undiagnosed diabetes mellitus and pre- diabetes were identified by fasting plasma glucose and OGTT. In this study, the cut-off points of fasting capillary blood glucose for detecting undiagnosed diabetes and pre-diabetes were evaluated, using receiver operator characteristic curve(ROC). Results Of 1351 subjects that having had oral glucose tolerance test, 230 cases were diagnosed as diabetes mellitus(7.3%), 166 cases(5.2%)as IFG, and 204(6.7%)as IGT under fasting capillary blood glucose as test variable and state variables according to the following criteria.(1)FPG≥7.0 mmol/L or/and 2hPG≥11.1 mmol/L(2)FPG<5.6 mmol/L (3)FPG<7.0 mmol/L and 7.8 mmol/L≤2hPG≤ 11.1 mmol/L, areas under three ROC curves were 0.905, 0.633 and 0.719, respectively. The cut-offvalues of screening for undiagnosed DM, IGT and IFG were 6.0 mmol/L, 5.7 mmol/L, and 5.7 mmol/L, respectively. When cut-off value of screening for undiagnosed DM was 6.0 mmol/L, the maximal sensitivity was 78.0% and specificity was 89.3%.But there were both lower sensitivity and specificity in screening for IFG and IGT according to the best predicting value(5.7 mmol/L)from the ROC curves(50.3% and 28.0% vs. 60.8% and 28.0%). Conclusion Fasting capillary blood glucose with the lower cut-point of 6.0 mmol/L in screening for undiagnosed diabetes mellitus alone, was relatively reliable, whereas for both IFG and IGT the fasting fingertip blood glucose tests were fallible. It was convenient and could be used in screening the DM at the community level.

Fingerprints of lipophilic components in the roots of Salvia miltiorrhiza and S.yunnanensis were analyzed by UPLC-DADand UPLC coupled with mass spectroscopy to evaluate the differences and similarities of the lipophilic components in the two kinds of herbs.The UPLC analysis of 18 batches of S.miltiorrhiza and 16 batches of S.yunnanensis was performed on a 25℃Thermo Accucore C_(18)column(2.1 mm×100 mm,2.6μm)by Shimadzu LC-20AD;mobile phase was 0.026%phosphoric acid(A)-acetonitrile(B)with gradient elution;flow rate was 0.4 m L·min~(-1);detection wavelength was set at 270 nm;injection volume was 2μL.The molecular structures of the lipophilic components were analyzed on a 25℃Thermo Accucore C_(18)column(2.1 mm×100 mm,2.6μm)by Thermo U3000 UPLC Q Exactive Orbitrap LC-MS/MS with a mobile phaseconsisting of 0.1%formic acid water(A)and 0.1%formic acidacetonitrile(B).The mass spectrometry was acquired in positive modes using ESI.There are 10 common peaks in the lipophilic components of S.miltiorrhiza.The similarity between the 16 batches of S.miltiorrhiza and their own reference spectra was greater than 0.942,and the average similarity was 0.973.There are 12 common peaks in the lipophilic components of S.yunnanensis.The similarity between the 18 batches of S.yunnanensis and their own reference spectra was greater than 0.937,and the average similarity was 0.976.The similarity between the reference chromatograms of S.miltiorrhiza and S.yunnanensis was only 0.900.There are three lipophilic components in S.yunnanensis,which are not found in S.miltiorrhiza,and one of which isα-lapachone.There is a lipophilic component in S.miltiorrhiza not found in S.yunnanensis,which may be miltirone.The two herbs contain 8 common lipophilic components including dihydrotanshinoneⅠ,cryptotanshinone,tanshinoneⅠ,tanshinoneⅡ_A,nortanshinone in which the content of tanshinoneⅡ_A,dihydrotanshinoneⅠand cryptotanshinone of S.yunnanensisis significantly lower than that of S.miltiorrhiza(P<0.01),and the contents of tanshinoneⅠand nortanshinone are significantly lower than that of S.miltiorrhiza too(P<0.05).There are significant differences in the types and contents of lipophilic components between the roots of S.miltiorrhiza and S.yunnanensis,and the similarity between the fingerprints of interspecies is much lower than that between the same species.Therefore,the roots of S.miltiorrhiza and S.yunnanensis are two kinds of herbs which are quite different in chemical compounds and compositions.
Chromatography, Liquid ; Diterpenes, Abietane ; Molecular Structure ; Plant Roots ; Salvia miltiorrhiza ; Tandem Mass Spectrometry

OBJECTIVE To provide certain therapeutic ideas and references for the pharmaceutical care of severe Legionella pneumonia in anti-infection treatment. METHODS Clinical pharmacists participated in the entire treatment process of a patient with severe Legionella pneumonia， and assisted clinical physicians in evaluating the infecting pathogens using the WUH （Winthrop- University Hospital criteria） scoring system， based on the patient’s clinical symptoms， physical signs， and changes in pulmonary imaging. Leveraging their pharmaceutical expertise， clinical pharmacists recommended a combination of piperacillin sodium and tazobactam with moxifloxacin hydrochloride for anti-infection treatment， and closely monitored the patient’s clinical manifestations. They promptly identified delirium and abnormally elevated levels of lipase， amylase and liver enzymes， and successively suggested adjusting the treatment plan to a combination of piperacillin sodium and tazobactam with doxycycline or azithromycin for anti- infection after analyzing the causes， along with liver protection treatment， enteral nutrition， and parenteral nutrition. Additionally， clinical pharmacists closely monitor the patient’s medication adherence and provide her with medication education. RESULTS The clinical physicians accepted the recommendations of the clinical pharmacists， and the patient improved after treatment and was discharged. A follow-up examination one month later showed no recurrence. CONCLUSIONS Clinical pharmacists， when assisting clinicians in treating severe Legionella pneumonia， not only pay attention to changes in the patient’s clinical symptoms and physical signs， but also closely monitor the adverse reactions of fluoroquinolone， tetracycline， and macrolide antibiotics. They should promptly recognize adverse reactions and provide recommendations for adjusting treatment plans， as well as offer comprehensive pharmaceutical care throughout the patient’s treatment， to ensure the effectiveness and safety of clinical therapy.