0.05.[Conclusion]VDR TruⅠ and FokⅠpolymorphisms are not related to LDD in Han nationality.

OBJECTIVE: To determine the changes of the plasma metabolic phenotype in rats with chronic restraint stress (rats with syndrome of liver qi stagnation and spleen deficiency), so as to reveal the biological features of the syndrome of liver qi stagnation and spleen deficiency, and to examine potential application of metabonomic analysis in studies of syndromes in traditional Chinese medicine. METHODS: Twenty-four male Sprague-Dawley rats were randomly divided into four groups: group A, 7 d normal control group; group B, 21 d normal control group; group C, 7 d stress group; and group D, 21 d stress group, with 6 rats in each group. Chronic restraint was used to induce stress in rats. Blood was collected from the cardio-ventricle under anesthesia on the 8th (groups A and C) or 22nd day (groups B and D) and detected by using the Fourier variable superconducting nuclear magnetic resonance (NMR) spectrometer (Varian UnityInova 600 M). Free induction decay signals were transferred into one-dimensional NMR spectrogram via 32 k Fourier transformation. Segmental integral calculus (0.04 ppm per segment) was performed from 4.5-0.5 ppm (Carr-Purcell-Meiboom-Gill, CPMG) or 6.0-0 ppm (longitudinal eddy-delay, LED) as defaulted 1H spectra values by using the VNMR software. Data were saved as text or excel files after normalization and then used for pattern recognition analyses. All the data were analyzed by principal component analysis (PCA) using the SIMCA-P 10.0 software (Umetrics AB, Umea, Sweden). RESULTS: The PCA analysis of rat plasma (1)H NMR spectra revealed different metabolic spectra between stress and control groups, which were consistent with alterations of in vivo metabolisms in rats under stress stimuli. Compared with the normal control group, rats with repeated stress displayed significant changes in spectral peak shapes of acetate, lactate, tyrosine, low-density lipoprotein, and unknown compounds (3.44 ppm). These altered metabolites can be used as biomarkers of syndrome of liver qi stagnation and spleen deficiency for further studies. CONCLUSION: The (1)H NMR spectra of metabolites in the rat blood are differentially changed after chronic stress. Specific, characteristic metabolic products can be identified by analyses of metabonomics, which lead to interpretation of biological feature of Chinese medicine syndromes. Therefore, metabonomic analysis is an approach with good development prospects to studies of TCM syndromes.

A PCR method was used to amplify the sequence encoding the mature peptide of?-mannanase of Bacillus subtilis. The gene was inserted into the Pichia pastoris vector pPIC9K, downstream of?-factor signal peptide sequence. The resultant recombinant plasmid pPIC9K-MAN was lineared by BglII digestion and introduced into the host Pichia pastoris GS115 by PEG method. After screen, the recombinant P. pastoris strain MAN22 was obtained and fermented in large scale 5L fermenter. The recombinant mannanase activity could reach to 1102IU/ml. The properties of the recombinant mannanase were characterized.

Objective To evaluate clinical efficacy of multiple regimen combination in treatment of osteoporosis of perimenopausal or postmenopausal women. Methods From Jul. 2008 to Dec. 2009, 109 women with low bone mineral density (BMD) or osteoporosis treated in Department of Obstetrics and Gynecology, Affiliated Second Hospital, Wenzhou Medical College were enrolled randomly into 3 group,including 36 women in Group A managed by osteoform 1000 mg/d + alfacalcidol 0. 25 μg/bid orally, 40 women in group B managed by osteoform 1000 mg/d + alfacalcidol 0. 25 μg/bid + tibolone 1.25 mg/d orally and 33 women in group C managed by ostcoform 1000 mg/d + alfacalcidol 0. 25 μg/bid +bisphosphonates 70 mg/w orally. After 48 weeks BMD on lumbar 1 -4 (L1-4) and left femur were detected by X-ray. Bone alkaline phosphatase(BALP) ,cross linked clelopeptide of type Ⅰ collagen(CTX) and 25-hydroxychole calciferol [25 (OH) D3] was measured by enzyme linked immunosorbent assay (ELISA).Result Seven women (6. 4%, 7/109) were withdrawed form this study, including 2 cases losing follow up in group A, 3 cases stopping treatment in group B, 2 cases giving up treatment due to severe adverse effect (burning in upper abdomen) in group C. (1) Pain relieve: after 48 weeks treatment, women in 3 groups improved symptom of pain significantly, the rates of pain relieve were 85% (29/34)in group A, 92% (34/37) in group B and 94% (29/31) in group C. (2) BMD: BMD was improved significantly in women in 3 groups after treatment. BMD of L1-4 were (0.88±0.15) g/cm2 in group A,(0.89±0.18) g/cm2 in group B and (0.87±0.10) g/cm2 in group C before treatment, and converted to (0.90±0.01) g/cm2 in group A, (0.93±0.09) g/cm2 in group B and (0.91±0.11) g/cm2 in group C after treatment. BMD of left femur were (0.87±0.07) g/cm2 in group A, (0.87±0.07) g/cm2 in group B and (0.85±0. 12) g/cm2 in group C before treatment and converted to (0.90± 0.03) g/cm2 in group A, (0.91±0.08) g/cm2 in group B and (0.89 ±0.12) g/cm2 in group C after treatment. It was shown significantly different BMD between group B or C and group A (P < 0. 01), however, there was no significant different BMD between group B and C (P >0. 05). (3) Index of bone metabolism: BALP were (26±6) μg/L in group A, (26±9) μg/L in group B and (28±7) μg/L in group C before treatment and converted to (22±5) μg/L in group A, (20±9)μg/L in group B and (22±8)μg/L in group C after treatment, which showed statistical difference (P < 0.05). CTX were (0.85±0.20) ng/L in group A, (0.84±0.47) ng/L in group B, and (0. 88 ±0. 11) ng/L in group C before treatment and converted to (0. 81 ±0. 19) ng/L in group A, (0. 77±0.33) ng/L in group B, and (0.82 ±0. 14) ng/L in group C after treatment, which showed statistical difference (P < 0. 05). Conclusions Those 3 regimens combination could be used in treatment of osteoporosis by decreasing bone conversion, increasing bone density, decreasing bone absorption. Regimen A was only suitable for basic therapy,the other two regimens could provide better treatment.

Objective To investigate the expression and significance of JWA in Acute Myeloid Leukemia (AML). Methods Bone marrow mononuclear cell specimens were taken from 22 AML patients in newly diagnosis stage and complete remission stage respectively , and the JWA expression were detected at RNA and protein level. Results (1) JWA was expressed in all the samples at RNA and protein level. (2) At protein level, JWA expression was higher in the cells from newly diagnosed AML patients than in those from patients in complete remission stage (P < 0.05). (3)The complete remission rate of patients with higher expression level of JWA (87.5%) was similar to those with lower expression (83.3%). The complete remission rate of patients with higher expression level of JWA after the first course (31.25%) was lower than those with lower expression (66.7%). The early recurrence rate of patients with higher expression level of JWA (42.86%) was higher than those with lower expression (20%). Conclusion JWA may play an important role in the development and progression of AML. Increased expression of JWA may be one of the causes of refractory and relapsed AML.

Objective To compare the efficacy of different therapies for idiopathic membranous nephropathy (IMN) patients, and to discuss their rationality. Methods The clinicopathological data and therapies of 76 patients with IMN in our hospital was retrospectively analyzed and reviewed, and the efficacy was followed up.According to the different therapies, 76 patients were divided into 4 groups, including symptomatic treatment group, glucocorticoid-alone group, immunosuppressant-alone group, and glucocorticoid in combination with immunosuppressant group (combination group). Comparison and analysis of the efficacy of the different therapies were made. Results (1) The incidence of nephrotic syndrome and 24-hour proteinuria of patients in symptomatic treatment group were significantly lower than those in glucocorticoid-alone group and combination group. (2) The remission rates of 4 groups were 56.3%,73.7%,66.7% and 78.9%, respectively. In general, no statistical differences were observed in the remission rates of patients among the symptomatic treatment group, glucocorticoid-alone group and combination group. The 2-year and 5-year renal survival rates were 89.2% and 79.3%, respectively. (3) Patients in glucocorticoid-alone group and combination group were divided into low-risk, moderate-risk and high-risk patients. No difference in remission rate was observed between the two therapies for low-risk and moderate-risk patients.But for high-risk patients,the remission rate in combination group was significantly higher than that in glucocorticoid-alone group.(4) Patients in glucocorticoid-alone group and combination group were divided into remission subgroup and non-remission subgroup. It showed that only estimated glomerular filtration rate (eGFR) between these two subgroups had statistical difference, and eGFR in non-remission subgroup was lower than that in the remission subgroup. Conclusions For high-risk patients,treatment with glucocorticoid combined with immunosuppressant may improve the remission rate of proteinuria significantly.Glomerular filtration rate before treatments is an important prognostic factor.

Objective To explore the role of brief ischemia pretreatment in the induction of renal ischemic tolerance,and investigate its effects on tubular cell necrosis,apoptosis and proliferation. Methods Male Sprague-Dawley rats were randomly divided into three groups,including sham-operated group (Sham),ischemia/reperfusion injured group subjected to theocclusion of both renal pedicles for 40 min followed by reperfusion(I/R),and preconditioned group with 20-min ischemia pretreatment induced 4 days before I/R(IPC).Histological changes were evaluated by PAS staining.The ultra-structure of tubular cells was observed by transmission electron microscopy(TEM).Apoptosis was confirmed by terminal deoxynucleotidyl transferase (TdT)-mediated dUTP-biotin nick end labeling (TUNEL).The proliferation of tubular cells was evaluated with proliferating cell nuclear antigen(PCNA). Results Twenty-minites ischemia pretreatment offered both promising functional and histological protection against 40-min ischemia/reperfusion injury (P<0.01).The mortality rate wag reduced from 33%in I/R group to 0 in IPC group.The renopmtection offered by 20-min ischemia pretreatment was accompanied with reduced postischemic tubular cell apoptosis and necrosis (P<0.05), and increased cell proliferation (PCNA positive) (P< 0.01). Conclusions Brief and sublethal prior ischemia can render the kidney more tolerant to subsequent prolonged I/R injury. Its ability to tilt the balance of tubular cell fate toward survival, reducing postischemic cell death and enhancing cell proliferation, may play an important role in renal protection of ischemic preconditioning.

Objective To investigate the alteration of mast cell(MC)in elderly patients with reflux esophagitis(RE). Methods Twenty eight elderly and 15 non-elderly patients with RE were recruited.Lower esophageal mucosal mast cells and the percentage of degranulated mast cells were countered after immunohistochemieal staining.The ultrastrueture of mast cells was observed by eleetromieroscope. Results The number of mast cells in lower esophageal mucosa in elderly patients with RE was significantly more than that in non-elderly patients with RE(10.24±2.56 VS.5.07±0.18,P＜0.01),and the percentage of degranulated mast cells in lower esophageal mucosa was also significantly higher in elderly patients with RE than in non-elderly patients with RE[(24.7±4.6)%vs.(13.5±5.5)%,P＜0.01].The more severe the esophageal mucosallesions,the more the numberof mast cells. Under the electromicroscopy, more Golgi apparatus, mitochondria and endoplasm-reticulum were found in mast cells.Special secreting particles were also found in cytoplasm,more vacuole were left behind after mast cells degranulation in elderly patients with RE.Conclusions Increased numbers of mast cells and mast cell activation may be involved in the pathogenesis of elderly RE.

Objective:To explore the efifcacy and safety for radiofrequency catheter ablation (RFCA) of left bundle branch guided by left bundle potential (LBP), X-ray image with EnSiteNavX System in canine model.
Methods:The RFCA of left bundle branch was conducted in 13 dogs. A mapping catheter was positioned in right atrium to record right-sided His-bundle (R-His) potential, and an ablation catheter via right femoral artery was retrograded to left ventriclefor LBP mapping and ablation. Meanwhile, EnSiteNavX System was used to identify R-His, L-His and LBP at the same time. The potential characteristics in dogs with successful ablation were observed, the PR interval, QRS shape and time limit, AH interval, HV interval, the A/V electro-gram ratio in ablationcatheter at before and after ablation were recorded. The procedural time and X-ray exposure time between LBP with X-ray image method and LBP, X-ray image with EnSiteNavX System method were compared.
Results: There were 9/13 dogs received successful left bundle branch ablation, 3 dogs failed and 1 suffered from complete A-V block. At the successful ablation target site, the LBP-V was (17.8 ± 2.6) ms with the range of (13-21) ms, and the A/V electro-gram ratio<1/10. The procedural time and X-ray expose time were signiifcantly decreased in LBP, X-ray image with EnSiteNavX System method than those in LBP with X-ray image method P=0.007 and P<0.001.
Conclusion:Under the LBP, X-ray image with EnSiteNavX System guidance method, left bundle branch could be safely and effectively ablated to establish left bundle branch block (LBBB) model in experimental canine.

Background and purpose: Increasing evidence has indicated that polymorphisms in the microRNA (miRNA, miR) binding site of target gene can alter the ability of miRNA and modulate the risk of cancer. We aimed to investigate the association between a miR-520a binding site single nucleotide polymorphism (SNP) rs141178472 in the PIK3CA 3’UTR and the risk of colorectal cancer in a Chinese Han population. Methods:The polymorphism rs141178472 was analyzed in a case-control study, including 386 colorectal cancer patients and 394 age-and sex-matched controls. The relationship between the polymorphism and the risk of colorectal cancer was examined by statistical methods. Results:Individuals carrying the rs141178472 CC genotype or C allele had an increased risk of developing colorectal cancer (CC vs TT, OR=1.716, 95%CI:1.084-2.716, P=0.022;C vs T, OR=1.258, 95%CI:1.021-1.551, P=0.033). Furthermore, the expression of PIK3CA was detected in the peripheral blood mononucleated cell of colorectal cancer patients, suggesting that mRNA levels of PIK3CA might be associated with SNP rs141178472. Conclusion:These ifndings provide evidence that a miR-520a binding site polymorphism rs141178472 in the PIK3CA 3’UTR may play crucial roles in the etiology of colorectal cancer.