Objective To detection the methylation of p16INK4A in primary hepatocellular carcinoma, a nested bisulfite sequencing and methylation-specific polymerase chain reaction (BS-MSP) protocol was designed and used.Methods Bisulfite-modified DNA were amplified to evaluate the quality of templates with a pair of bisulfite sequencing primers in the first round of PCR, then subjected to methylation assay with corresponding methylation or unmethylation specific PCR primers.Representative PCR products were sequenced to confirm its correctness.Results 3 of 40 cases (7.5%) were failed to assay due to poor quality of templates, and 29 of 37 cases (78%) were detected p16INK4A methylation.Sequencing results confirmed that templates were correctly amplified.Conclusion BS-MSP technique might be valuable for methylation study on carcinogenesis and clinical assay.

p70 ribosomal protein S6 kinase (p70S6K), an important member of AGC family, is a kind of multifunctional Ser/Thr kinases, which plays an important role in mTOR signaling cascade. The p70 ribosomal protein S6 kinase is closely associated with diverse cellular processes such as protein synthesis, mRNA processing, glucose homeostasis, cell growth and apoptosis. Recent studies have highlighted the important role of S6K in cancer, which arose interests of scientific researchers for the design and discovery of anti-cancer agents. Herein, the mechanisms of S6K and available inhibitors are reviewed.
Antineoplastic Agents ; Humans ; Protein Kinase Inhibitors ; chemistry ; Ribosomal Protein S6 Kinases, 70-kDa ; antagonists & inhibitors ; metabolism ; Signal Transduction ; TOR Serine-Threonine Kinases

In this study, we use pot experiment to evaluate the effect of plant growth regulator on plant morphology and biomass allocation of Salvia miltiorrhiza. Different concentrations of uniconazole were supplied to S. miltioohiza by means of foliar spray. Height, breadth and stem diameter were measured dynamically, the biomass of leaf, stem, flower and fruit, root biomass and biomass ratio were also examined at the harvest time. Owing to the treatment, plant morphology showed significant changes, the height had been greatly reduced and the breadth decreased largely. Meanwhile, the biomass allocation changed too. The biomass ratio of leaf and stem had been notably reduced while the biomass ratio of root had been increased remarkably. It appears that foliar application of uniconazole during vigorous growth period in S. miltioohiza has dramatic effect on dwarfing plant and improving resistant to lodging. This measure could also be applied to condensed cultivation of S. miltioohiza to increase production.
Biomass ; Plant Growth Regulators ; pharmacology ; Plant Leaves ; drug effects ; growth & development ; Plant Roots ; drug effects ; growth & development ; Plant Stems ; drug effects ; growth & development ; Salvia miltiorrhiza ; drug effects ; growth & development ; Triazoles ; pharmacology

Background In recent years,with the contiunous progress of the refractive surgery,the operation skill of phakic intraocular lens(PIOL)implantation for correcting extreme high myopia,astigmatism,farsightedness have made greater progression,and its security,effectiveness in clinical attract much more attention. Objective This study was to evaluate the efficacy,safety and stability of Toric Implantable Collamer Lens(TICL)for extreme high myopic astigmatism. Methods This retrospective case series included 33 eyes of 27 patients from May 2008 to February 2009.A TICL was intraocularly implanted via a 3 mm clear corneal incision after paraocular anesthesia.Patients were examined preoperatively and followed-up at 1 day,1 week,1 month,3,6,12 and 18 months postoperatively.The examinations included uncorrected visual acuity,best corrected visual acuity(BCVA),slit lamp examination,refraction,intraocular pressure,endothelial cell morphometry,etc.The written informed consent was obtained from each patient before any medical procedure. Results The uncorrected visual acuity in 96.97% eyes was equal or improved after operation in comparison with BCVA of preoperation.The spherical refraction was within-1.00 D-+0.25 D.The cylinder refraction was within-1.00 D-0 D.The axial deviation of TICL within 10 degree was 93.94%(31/33).No significant differences were found in the intraocular pressure and endothelial cell morphometry between preoperation and postoperation(intraocular pressure:F=3.35,P=5.49;endothelial cell morphometry:t=1.835,P=0.082).The visual acuity and refraction were stable during the follow-up.Astigmatic axial rotation required surgical intervention on one eye.One eye occurred high intraocular pressure because of bigger TICL diameter.The intraocular pressure returned to normal after TICL was exchanged.No cataract occurred during the follow-up duration. Conclusion TICL implantation appears to be an effective,safe and reliable method for extreme high myopic astigmatism.

Objective:This study aimed to detect the special methylation profile in peripheral blood for hepatocellular carcinoma (HCC). Methods:The methylation status of 12 tumor suppressor genes (TSGs) in the plasma of 55 HCCs and 54 chronic liver diseases (CLDs) was tested by methylation-specific PCR (MSP). Results:In HCC, the methylation frequencies were 78.18%in APC, 63.64%in cyclin D2, 58.18% in TFPI2, 49.09% in DKK3, 49.09% in GSTP1, 47.27% in p16, 40.00% in Sigma 14-3-3, 18.18% in SFRP2, 16.36% in ppENK, 9.09% in DKK2, 7.27% in NPTX2, and 5.45% in LHX1. In CLD, the methylation frequencies were 27.78% in APC, 22.22%in cyclin D2, 7.41%in TFPI2, 3.70%in DKK3, 16.67%in GSTP1, 37.04%in p16, 37.04%in Sigma 14-3-3, 11.11%in SFRP2, 20.37%in ppENK, 7.41%in DKK2, 7.41%in NPTX2, and 9.26%in LHX1. The methylation frequencies of APC, cyclin D2, TFPI2, DKK3, and GSTP1 were higher in HCC than in CLD (P<0.01). The methylation index (MI) of the five-gene methylation profile was statistically higher in HCC (median, 0.6;IQR, 0.4-0.8) than CLD (median, 0.2;IQR, 0-0.2) (P<0.01). In HCC, MI was statistically related to the patient's age. Older patients with HCC had a higher MI. No significant correlation was observed between MI and other clinicopathological data. Moreover, MI was not related to the disease free survival and the overall survival in HCC. Conclusion:This five-gene methylation profile may be a promising biomarker for the assistant diagnosis of HCC.

Objective To report the clinical and neuroimaging features in three cases of neurosyphilis with mesiotemporal lobe lesions; and to discuss possible pathogenesis and differential diagnosis.Methods The clinical manifestations,laboratory examinations,neuroimaging features,differential diagnosis and curative effect of 3 cases of neurosyphilis with mesiotemporal lobe lesions were analyzed retrospectively.The possible pathogenesis was discussed.Results All of 3 cases had positive and high titer index of syphilis in both serum and cerebrospinal fluid.The onset symptom was abnormal cognitive functions with acute aggravation or rapid progression.Cerebrospinal fluid analysis implied lymphocyte predominant inflammatory reaction with increased protein.Electroencephalography showed localized slow waves as to the lesions.On brain magnetic resonance imaging,unilateral or asymmetrical bilateral mesiotemporal lobe was affected with long T2 signal.Herpes simplex virus encephalitis and paraneoplastic limbic encephalitis should beconsidered in differential diagnosis.The symptoms and laboratory indexes improved significantly after anti-syphilis treatment.Conclusions Neurosyphilis should be evaluatedin a patient with onset of abnormal cognitive functions and having mesiotemporal lobe lesions on magnetic resonance imaging.Promptly diagnosis and early treatment could achieve good prognosis.

Objective To investigate the levels of serum Golgi protein(GP73) (sGP73) in patients with HBV-related liver disease and investigate the role of sGP73 as an indicator for diagnosis of hepatocelluar carcinoma (HCC).Methods The concentration of sGP73 in patients with chronic hepatitis B (CHB,n =31),liver cirrhosis (LC,n =60),HCC (n =71),self-limited HBV infectors (n=21 ) and healthy controls (n =42) were tested by enzyme-linked immunosorbent assay (ELISA) assay and statistically analyzed in combination with relevant clinical indicators.Results The median of sGP73 in HBV-related liver disease group was significantly higher than that in the groups of self-limited HBV infectors and healthy controls respectively (P＜0.01).Among the groups with HBV-related liver disease,the median of sGP73 in LC group (231.13 ng/ml) was significantly higher than that in HCC without treatment group ( 117.63 ng/ml) (P ＜ 0.01 ) and CHB group (93.09 ng/ml) (P＜0.01).No significant difference was shown between HCC (without treatment) group and CHB group (P＞ 0.05),neither between self limited HBV infectors and healthy controls (respectively,36.79 ng/ml and 45.40 ng/ml) (P ＞ 0.05). The median of sGP73 in post-operation group (175.12 ng/ml,n=52) was significantly higher than in pre-operation HCC group (107.28 ng/ml,n=52) (P＜0.01).Along with the decreasing of liver function,sGP73 level was elevated in groups with HCC or LC.The receiver operating curve (ROC) constructed with the ratio of AFP and GP73 (AFP/GP73) showed a sensitivity of 78.87 ％ and specificity of 86.21 ％ with an area under the receiver operating curve (AUROC) of 0.878 (95％ CI:0.817-0.938) for diagnosis of HCC; comparably,a sensitivity of 67.61％ and specificity of 85.12％ were shown with a AUROC of 0.826 (95％ CI:0.755-0.897) when performed with AFP.Conclusion The level of sGP73 in HBV-related liver disease group is higher than that in the groups of self-limited HBV infector and healthy control,and it is positively correlated with the degree of hepatic impairment.For the diagnosis of HCC,joint detection of AFP and GP73 could achieve a better combination of sensitivity and specificity than the independent AFP test.

ObjectiveTo investigate hepatocyte growth factor (HGF) levels in the tissue and serum of patients with chronic hepatitis,cirrhosis or hepatocellular carcinoma (HCC),and analyze the clinical significances of HGF for HCC.MethodSurgical specimens from 97 patients were collected during Dec.2003 to Aug.2008 in the Third Central Hospital.The patients were prospectively enrolled and categorized into four groups:normal subjects ( n =11 ),chronic hepatitis B or C ( n =6＝,cirrhosis ( n =20)and HCC ( n =60 ) including well-differentiated ( n =21 ),moderately differentiated ( n =23 ),poorly differentiated (n =16) specimens.N0 (n =24),N1 (n =21 ),N2 (n =54) and N3 (n =43) were tissues respectively removed from liver at 0,1,2 or 3 cm beyond the margin of tumor.HGF mRNA expression in liver tissues was determined by real-time quantitative reverse transcription- (RT)-PCR.Serum HGF levels in the other cases of normal subjects ( n =20),chronic hepatitis B or C ( n =20),cirrhosis ( n =20) and HCC (n =57) were measured by ELISA.The Kaplan-Meier method with log-rank test was employed for survival analysis.Univariate and multivariate analyses were performed to identify prognostic factors in each group.ResultsThe HGF mRNA in normal subjects,chronic hepatitis,cirrhosis,N3,N2,N1,N0 and HCC were0.99(0.78-1.66),2.15(1.06-3.40),1.78(1.18-2.73),4.59(2.67 -8.63),3.86 ( 2.25 - 6.45 ),3.12 ( 1.59 - 5.74 ),2.92 ( 0.88 - 5.99 ) and 0.48 ( 0.19 - 1.06 ) respectively.The serum concentration of HGF in the normal subjects,chronic hepatitis,cirrhosis and HCC patients were (0.31 ± 0.05 ),(0.65 ± 0.07 ),( 1.27 ± 0.30 ) and ( 2.06 ± 0.66) μg/L respectively.The highest level of HGF mRNA was found in N3,while the HGF mRNA expression in HCC was [ (2.14 ± 0.52 ) μg/L] lower than that not only in the non-tumor tissues,but also in the normal control ( U =196.50,P =0.03 ).The serum concentration of HGF was significantly higher in patients with chronic hepatitis,cirrhosis or HCC than in normal subjects.The serum HGF level of HCC was bounced after hepatectomy (t =2.70,P ＜0.01 ).On the logistic regression analysis,the tumor numbers and Child-pugh were related with the levels of the tissue HGF mRNA and serum HGF of HCC,OR were0.15 (95％CI:0.03-0.72,P＜0.05) and0.13 (95％CI:0.27 -0.89,P ＜0.05 ),respectively.Univariate analysis using the Cox proportional hazards model in the complication groups revealed that the levels of the tissue HGF mRNA and serum HGF were significant risk factors of death for HCC,OR were 0.02 (95％ CI:0.00 - 0.52,P ＜ 0.05 ) and 10.01 (95％ CI:1.16 -86.23,P ＜ 0.05 ),respectively.On the Log-rank analysis,no statistically difference in the cumulative survival was found between the two groups categorized by median (0.49) of tissue HGF mRNA 2 - AACT (X2 =0.13,P =0.72).While the HCC patients were dichotomized by their the median(0.69 μg/L) of serum HGF concentration,the death risk for the patients with higher levels of HGF was increased 2.84 fold than those with lower levels (95％ CI:1.03 - 7.92,P ＜ 0.05 ).ConclusionHGF mRNA expression is decreased in tumor tissues,while its level in tumor adjacent live and serum is significantly elevated and is in association with shortened postoperative survival of HCC patients.

AIMIn order to elucidate the underlying mechanism of depressed maximal isometric twitch tension normalized by cross sectional area of muscle strip in unloaded soleus.

METHODSThe soleus and extensor digitorum longus (EDL) muscle strips were perfused in vitro and treated by 2,3-Butanedione monoxime (BDM).

RESULTSThe BDM decreased Pt of soleus and EDL in a concentration-dependent manner. The Pt could restored completely to normal level after washing out BDM. The isometric twitch duration was not altered during 1 mmol/L BDM of perfusion, but was shortened at 10 mmol/L dose. The time from maximal to half Pt in EDL was shorter than that in soleus during 10 mmol/L BDM of perfusion. The inhibitory effects of BDM on myosin ATPase activity were higher in EDL than in soleus. The inhibitory extent of BDM on myosin ATPase activity of soleus and EDL was lower than that on Pt.

CONCLUSIONThese results suggest that reduction in cross-bridge function of skeletal muscle may be one of reasons induced a decrease in its Pt. BDM is not a specific inhibitor on myosin ATPase activity and can affect multiple parts of excitation-contraction coupling in skeletal muscle.

Animals ; Diacetyl ; analogs & derivatives ; pharmacology ; Isometric Contraction ; drug effects ; physiology ; Male ; Muscle Contraction ; drug effects ; physiology ; Muscle, Skeletal ; drug effects ; physiology ; Myosins ; antagonists & inhibitors ; Rats ; Rats, Sprague-Dawley

The present study was performed to observe the time course and features of intermittent tetanic contractile function changes in soleus and extensor digitorum longus (EDL) muscles of tail-suspended rats. The optimal stimulating frequency, fatigability and time-dependent recovery after fatigue were measured in isolated muscle strips. The optimal stimulating frequency of soleus and EDL was 60 Hz and 120 Hz in control rats, respectively. It was not changed in 1-week unloaded soleus, but shifted to 80 Hz and 100 Hz in 2- and 4-week unloaded soleus, respectively. The maximal isometric tension (P(o)) of tetanic contraction at optimal stimulating frequency did not alter in 1- and 2-week unloaded soleus, but significantly decreased in 4-week unloaded soleus. After 5 min of fatigue, tetanic contractile tension of control soleus was decreased to 22.8% P(o), but significantly decreased to 10.4%, 10.0% and 11.6% P(o) in 1-, 2- and 4-week unloaded soleus, respectively. The tetanic contractile tension recovered to 98% P(o) in control soleus at the twentieth minute after fatigue, but only recovered to 79.0%, 83.6% and 78.5% P(o) in 1-, 2- and 4-week unloaded soleus. The optimal stimulating frequency, P(o), fatigability and time-dependent recovery of intermittent tetanic contraction were not altered in unloaded EDL compared with control. These results indicate that higher stimulating frequency can compensate the P(o) reduction in 1- and 2-week unloaded soleus, but not in 4-week unloaded soleus. The unloaded soleus, but not EDL, is more susceptible to fatigue than the synchronous controls. The unloaded soleus not only fatigues to a greater extent but also recovers significantly less than the control.
Animals ; Electric Stimulation ; In Vitro Techniques ; Male ; Muscle Contraction ; physiology ; Muscle Fatigue ; physiology ; Muscle, Skeletal ; physiology ; physiopathology ; Muscular Atrophy ; physiopathology ; Rats ; Rats, Sprague-Dawley ; Weightlessness