To study the constituents of Rubus corchorifolius L. f, the constituents were separated by using various chromatographies and the structures were identified on the basis of chemical and spectral analysis. Two compounds were isolated from Rubus corchorifolius L. f. Their structures were identified as ent-kauran-3beta, 16beta, 17, 19-tetrol (1), ent-2-carbonyl-16beta-hydroxy-kauran-17-beta-D-glucoside (2). Compounds 1 and 2 are new compounds.
Diterpenes, Kaurane ; chemistry ; isolation & purification ; Glucosides ; chemistry ; isolation & purification ; Molecular Structure ; Plant Leaves ; chemistry ; Plants, Medicinal ; chemistry ; Rosaceae ; chemistry

OBJECTIVEHemophilia A is an inherited bleeding disorder caused by defects in factor VIII (FVIII) gene. In the present study, the frequencies of the microsatellite alleles at introns 13 and 22 in the factor VIII gene were analyzed in the group of Han nationality in Guangxi Zhuang Autonomous Region to explore their diagnostic value for hemophilia A. These two sites were also used to detect the carriers in 13 hemophilia A families.

METHODSNinty-one individuals of Han ethnic group in Guangxi Zhuang Autonomous Region (135 X chromosomes) and 13 HA families were subjected to molecular studies. First, these two fragments were PCR amplified simultaneously. Then, silver staining was used later to show their polymorphisms. The investigators selected one sample at random to obtain its lengths of the PCR products at these two sites by ABI310 PCR amplifier. After counting its repeated numbers of (CA) according to the documents concerned, the repeated numbers of the other samples could be counted easily.

RESULTSIn the 91 individuals, 6 and 4 alleles were detected at these two sites, respectively. At intron 13 the allele frequencies ranged from 0.0002 to 0.5408 and polymorphism information content (PIC) was 0.5899. At intron 22 the allele frequencies ranged from 0.0444 to 0.4963 and its PIC was 0.5359. The actual heterozygosity for intron 13 and intron 22 were 0.6364 (28/44) and 0.5227 (23/44), respectively. In 13 hemophilia A families with positive history, 9 of them were diagnosed by this method and the diagnosis rate was 69%.

CONCLUSIONWith high PICs, (CA)n at intron 13 and intron 22 were two valuable sites in the diagnosis of hemophilia A in the population of Han ethnic group in Guangxi Zhuang Autonomous Region. Compared with some other HA restrictive fragment length polymorphisms (RFLP), intron 22 (GT)n (AG)n was more informative.

Alleles ; Amplified Fragment Length Polymorphism Analysis ; Asian Continental Ancestry Group ; genetics ; Factor VIII ; genetics ; Female ; Gene Frequency ; Genetic Predisposition to Disease ; Hemophilia A ; diagnosis ; genetics ; Heterozygote ; Humans ; Introns ; Male ; Polymerase Chain Reaction ; Polymorphism, Genetic ; Sequence Analysis, DNA ; Silver Staining

Objective To explore the siRNA as a new antiviral therapy,evaluate the inhibition effect of siRNA based on vector on the HBV of HepG2.2.15 cell,and observe the side effect and toxicity of siRNA vector on cells and the off-target effect of siRNA.Methods Three pairs of siRNA duplexes targeting HBV C gene were designed as double strands,and the duplex were annealed and ligated into the p-Silencer-Cmv 4.1-hygro vector.The ligation products were used to transform JM109 cells.The clones with shRNA were obtained,and the vectors were purified.After the initial identification of the vector with agarose gel and the size of the inserted sequence got examined by native polyacrylamide gel electrophoresis,furthermore the sequencing was further carried out.The recombinant plasmids were purified with ultrapure Midipreps DNA Purification System.Then HepG2.2.15 cells were transfected with the plasmid mixed with siPort XP-1.The expression of HBsAg and HBeAg were detected by immunofluorescence and Western blot,and the HBV RNA was investigated by RT-PCR.Furthermore the real-time quantitive PCR was carried out to detect the changes of HBV DNA.In order to evaluate the toxicity of the shRNA,MTT was used to examine the growth rate and curve of cells.The ELISA was performed to detect the changes of interferon-? (IFN-?).Results The Western blot showed that the HBsAg and HBeAg protein were suppressed with (81.15?0.69)%,(88.12?0.92)% respectively by vector p-C2 on the third day of post-transfection.It had the similar result indicated by immunofluorescence.And the RT-PCR showed that the specific siRNA targeting HBV C gene could markedly suppress the expression of HBV mRNA and the HBV C gene mRNA was inhibited with 96.9%.The real-time quantitive PCR showed that the specific functional siRNA could markedly suppress HBV DNA copy with two orders of magnitude,while the siRNA vector had no effect on the growth of cell showed by MTT detection.Compared with the non-transfected group and p-NC group,the IFN-? level was almost the same with siRNA p-C1,p-C2,p-C3 groups.Conclusions The siRNA based on the expression vector can suppress the expression and replication of HBV in HepG2.2.15 cell.The inhibition effect was specific and had a certain dependency on siRNA concentration.No toxicity effect was found in the study.And the drug resistance wouldn′t happen because the silence was based on the split of gene.

OBJECTIVETo investigate the effects of a high-carbohydrate diet on the lipid and apolipoprotein ratios in healthy young adults with different genotypes of the polymorphism at -75 site in the promoter region of the gene of apolipoprotein AI (APOA1).

METHODSFifty-six subjects aged (22.89 +/- 1.80) years were given a wash-out diet for 7 days, followed by a high-carbohydrate diet for 6 days. The wash-out diet contained 15% protein, 31% fat, and 54% carbohydrate. The high-carbohydrate diet contained 15% protein, 15% fat, and 70% carbohydrate. Twelve-hour fasting serum lipids and apolipoproteins B100 and AI were measured on the mornings of the 1st, the 8th, and the 14th days from the beginning of the wash-out diet. The ratios of triglyceride (TG)/high-density lipoprotein cholesterol (HDL-C), total cholesterol (TC)/HDL-C, low-density lipoprotein cholesterol (LDL-C)/HDL-C, and apolipoprotein B100 (APOB100)/apolipoprotein AI (APOAI) were calculated. The genome DNA was extracted and the polymorphism of APOA1 -75 G/A was determined by polymerase chain reaction followed by restriction fragment length polymorphism assay.

RESULTSAt baseline, the lipid and apolipoprotein ratios showed no significant differences between the GG genotype and the A carriers in males (P > 0.05), whereas the female A carriers had a significantly higher ratio of LDL-C/ HDL-C compared with the female subjects with the GG genotype (P < 0.05). Following the high-carbohydrate diet, significant decreases of TC/HDL-C were found in all the groups, regardless of sex and genotype (P < 0.01). LDL-C/HDL-C experienced significant decreases in both the genotypes in males (P < 0.05), while in females, significant decrease of LDL-C/HDL-C was only observed in A carriers (P < 0.01).

CONCLUSIONThe A allele of the -75 G/A polymorphism in APOA1 may have specific effects on the LDL-C/HDL-C ratio in females.

Adult ; Apolipoprotein A-I ; genetics ; Apolipoproteins ; blood ; Dietary Carbohydrates ; metabolism ; Female ; Genotype ; Humans ; Lipids ; blood ; Male ; Polymorphism, Genetic ; Young Adult

Objective:To compare the clinical efficacy of transperitoneal robot assisted laparoscopic radical prostatectomy (RARP) versus extraperitoneal single port RARP.Methods:A retrospective analysis was perfoumed on 142 cases of RARP from July 2019 to June 2020 in Robotic Minimally Invasive Center of Sichuan Provincial People's Hospital, including 82 cases(Group A) , aged 70.0(65.6-78.0), undergoing transperitoneal RARP and 60 cases(Group B), aged 68.2 years old(60.1-79.2), undergoing extraperitoneal single port RARP. For group A, PSA was 12.9(5.6-64.0) ng/ml, with 26 cases of less than 10 ng/ml(31.7%), 40 cases of 10-20 ng/ml(48.8%), and 16 cases of more than 20 ng/ml(19.5%), the Gleason score was 7.2(6.0-10.0), with 14 cases(17.1%) of ≤6, 56 cases(68.3%) of 7, and 12 cases(14.6%) of ≥8, and the prostate volume was 61.3(29.0-112.0) ml. There were 49 cases with BMI≤25 kg/m 2, accounting for 59.8%, and 33 cases with BMI>25 kg/m 2, accounting for 40.2%. There were 17 cases(20.7%) of T 1, 44 cases(53.7%) of T 2 and 21 cases(25.6%) of T 3. The proportion of lymph node dissection was 17.1%, and 4 cases(4.9%)had a history of operation. For group B, the PSA was 12.2(1.0-42.6)ng/ml, with 20 cases (33.3%) of <10 ng/ml, 31 cases(51.7%)of 10-20 ng/ml, and 9 cases (15%) of >20 ng/ml. Gleason score was 7.1(6.0-9.0), with 12 cases (20.0%) of ≤6, 42 cases (70.0%) of 7, and 6 cases (10.0%)of ≥8. Prostate volume was 42.4(31.2-72.8)ml on average. There were 37 cases (61.7%) with BMI≤25 kg/m 2, and 23 cases (38.3%)with BMI >25 kg/m 2 . There were 17 cases(28.3%)of T 1, 32 cases(53.3%)of T 2 and 11 cases(18.3%)of T 3.The proportion of lymph node dissection was 11.7% and 4 cases (6.7%) had a history of operation.There was no statistically significant difference in term of age, PSA level, Gleason score, BMI, clinical stage, proportion of lymph node dissection or history of operation between the two groups( P>0.05), but there was statistically significant difference for prostate volume( P<0.05). All operations were performed by the same operator. Four different ways of bladder neck and urethral dissociation was selected according to the intraoperative conditions in Group A, include VIP style, T-shape incision style, VIP plus T-shape incision style or the style along the lateral side of the bladder neck. Small and single anterograde incision stripping of bladder neck was routinely performed in the Group B. Postoperative follow-up was performed to compare the operation time, intraoperative blood loss, bladder neck and urethral anastomosis time, postoperative hospital stay, postoperative exhaust time, postoperative complications, positive rate of surgical margin, indwelling time of urinary catheter, urinary continence satisfaction rate of immediately after operation, 3 months and 6 months after operation, wound healing and aesthetics. Results:All of the operations were successfully completed under robot-assisted laparoscopy, and there was no conversion to open surgery. The operation time was 56.0(45.0-112.0) min in the Group A and 65.4(55.5-96.8) min in the Group B, and there was no statistically significant difference( P>0.05). The intraoperative blood loss was 76.2(30.0-120.5) ml and 55.6(45.5-114.6) ml, respectively, and the difference was not statistically significant( P>0.05). The time of bladder neck urethral anastomosis was 18.9(12.6-25.6) min and 16.2(10.7-19.3) min, respectively, and the difference was not statistically significant( P>0.05). The postoperative hospital stay days were 9.3(8.0-16.0) d and 8.4(7.0-13.0) d, respectively, and the difference was not statistically significant( P>0.05). The postoperative exhaust time was 1.3(0.7-3.0) d and 3.4(2.0-7.0) d, respectively, and the difference was statistically significant( P<0.05). There was 1 case of anastomotic fistula with ureteral injury in Group A, and no serious complication in Group B, and the difference was not statistically significant( P>0.05). The number of positive surgical margin in the two groups was 13(15.9%)and 9(15.0%)respectively, and the difference was not statistically significant( P>0.05). The indwelling time of urinary catheter after operation was 9(7-21) d and 6(4-8) d, respectively, and the difference was statistically significant( P<0.05). The number of patients with satisfactory urinary continence immediately after surgery, 3 months and 6 months after surgery in the two groups were 8(9.8%), 51(62.2%), 62(75.6%) and 17(28.3%), 43(71.7%) and 54(90.0%), respectively. The differences were statistically significant( P<0.05). The total incision lengths in the two groups were 12.1(10.4-13.4) cm and 5.6(5.0-6.0) cm, respectively, and the difference was statistically significant( P<0.05). Conclusions:The extraperitoneal single port RARP is safe and feasible, and the postoperative effect is similar to that of transperitoneal RARP. It has the advantages of shorter recovery time, higher urinary continence satisfaction rate, neater and more beautiful incision. The long-term therapeutic effect needs further confirming by prospective study.

AIMTo obtain Clenbuterol monoclonal antibodies.

METHODSClenbuterol complete antigen was prepared with diazotization method. BALB/c mice was immunized with subtractive immunization, Clenbuterol monoclonal antibody was prepared with rule hybridoma technique.

RESULTSThe mice obtained tolerance to BSA by subtractive immunization. The rate of the hybridoma cell with positive reaction which had obtained was 8.2%, and the specific clenbuterol monoclonal antibody was obtained at last.

CONCLUSIONMonoclonal antibodies to micromolecule contaminant be prepared by subtractive immunization, could decrease the workload in the bolting of monoclonal antibodies, and increase the chance to obtain the antibody of expected.

Animals ; Antibodies, Monoclonal ; biosynthesis ; immunology ; Clenbuterol ; immunology ; Female ; Hybridomas ; metabolism ; Immunization ; methods ; Male ; Mice ; Mice, Inbred BALB C ; Serum Albumin, Bovine ; immunology

The study was aimed to investigate the effects of total saponins of Panax notoginseng (tPNS) on cobalt chloride (CoCl2)-induced apoptosis of rat bone marrow mesenchymal stem cells (rBMSCs) and the underlying mechanism. rBMSCs were isolated by density gradient centrifugation from Sprague Dawley (SD) rats. After being incubated with different concentrations of tPNS (1, 10, 100 μg/mL) for 48 h, the rBMSCs were stained with EdU and PI for proliferation and cell cycle assay, respectively. CoCl2 group was treated with 300 μmol CoCl2 for 24 h, and different concentrations tPNS groups were treated with 300 μmol CoCl2 plus 1, 10 or 100 μg/mL tPNS. After Annexin V-FITC/PI staining, flow cytometry was applied to measure the cell apoptosis. For mitochondrial membrane potential assay, rhodamine123 and Hoechst33342 staining were used. qRT-PCR was applied to analyze gene expression of Bcl-2 family. The results showed that the proliferation rates of the three concentrations tPNS groups were all higher than that of the control group (all P < 0.05). Compared with control group, only 100 μg/mL tPNS group exhibited increased cell percentage of S and G2 phase. Compared with that in control group (without CoCl2), the apoptotic rate was increased by 14.2% in CoCl2 group. And the apoptotic rates were reduced by 14.4%, 12.8% and 13.9% in three concentrations tPNS groups, compared with that in CoCl2 group (all P < 0.01). CoCl2 could decrease the mitochondrial membrane potential, while different concentrations of tPNS reversed the inhibitory effect of CoCl2. Bcl-2 and Bcl-xl mRNA expressions in all tPNS groups were higher than those in CoCl2 group (all P < 0.05). Moreover, 10 and 100 μg/mL tPNS groups showed lower ratios of Bax/Bcl-2, compared with CoCl2 group. The results suggest that tPNS protects the rBMSCs against CoCl2-induced apoptosis through improving the cell mitochondrial membrane potential, up-regulating the expressions of anti-apoptosis genes Bcl-2 and Bcl-xl, and reducing the Bax/Bcl-2 gene expression ratio.
Animals ; Apoptosis ; Bone Marrow Cells ; Membrane Potential, Mitochondrial ; Mesenchymal Stromal Cells ; Mitochondria ; Panax notoginseng ; Rats ; Rats, Sprague-Dawley ; Saponins

Objective:To evaluate the feasibility and clinical efficiency of robot-assisted laparoscopic radical prostatectomy (RARP) via extraperitoneal PORT-free single incision approach.Methods:The data of 33 patients with prostate cancer underwent the extraperitoneal PORT-free single incision RARP from November 2020 to January 2021 in Sichuan Provincial People's Hospital was retrospectively reviewed. The average age was 66.7 (58-78) years, the median PSA was 20.77 (2.89, 56.44) ng/m, and the mean Gleason score was 7.0 (6.0-9.0). The mean prostate volume was 48.4 (25.0-220.0) ml. Clinical stage: 32 cases was in cT 2a-2cN 0M 0, 1 case in cT 3aN 0M 0. 16 cases had a history of operation. All 33 operations were performed by the same operator. All operations were performed by extraperitoneal PORT-free single-incision approach. The surgical condition, postoperative complication, pathology, and follow-up results were observed. Results:In this study, 33 operations were successfully completed without conversion to open or additional single hole channel instruments. The average operation time was 61.3 (38.0-120.0) min, with the mean intraoperative bleeding volume of 72.2 (45.0-220.0) ml and the mean bladder neck urethral anastomosis time of 11.7 (8.5-15.7) min. The mean postoperative hospital stay was 7.9 (6.0-15.0) d, the mean postoperative indwelling time of urinary catheter was 6.8 (6.0-14.0) d, and the mean postoperative evacuation time was 1.0 (0.5-3.0) d. The average incision length was 5.2 (4.6-5.8) cm. There was no obvious complications. The postoperative pathological stage: 21 cases were in < pT 3a, 12 cases were in ≥ pT 3a, and 6 cases (18.8%) had positive resection margin. 29 cases (88.9%) acquired satisfactory urinary continence after operation, and the frequency of urinary pad use was ≤ 1 tablet/day. Conclusions:The extraperitoneal single-incision RARP surgical channel without PORT is safe and feasible with a satisfying cosmetic effect, which saves costs and requires less specific channel device. Simultaneously, the new approach has strong replicability, short-term tumor control and urinary control effect with rapid postoperative recovery. However, the sample size of this study is relatively small, which needs further research and demonstration

OBJECTIVE: To explore the MRI findings of os acromiale and to analyze the relationship between os acromiale and the supraspinatus and infraspinatus injury. METHODS: From January 2010 to August 2020, 21 patients with os acromiale (os arcomiale group) were compared with 21 subjects with no evidence of os acromiale (no os arcomiale group). There were 14 males and 7 females in the os arcomiate group, aged from 29 to 77 years old, mean aged (55.5±11.5) years old. While in the control group, there were 10 males and 11 females in no os arcomiale group, aged from 31 to 70 years old, mean aged (51.1±10.0) years old. The os acromiales were classified as edematous os acromiale or non-edematous os acromiale based on whether the presence of marrow edema, and as displaced os acromiale or non-displaced os acromiale based on whether the presence of displacement of the os acromiale. The MRI features of os acromiale were analyzed. Statistical analyses were performed to identify the differences between the os arcomiale group and no os arcomiale group regarding rotator cuff tear, supraspinatus and infraspinatus injury. Differences in the supraspinatus and infraspinatus tear between the edematous and non-edematous os acromiale group, the displaced and non-displaced os acromiale group, the displaced os acromiale and no os arcomiale group were also assessed. RESULTS: On MRI, all the 21 os acromiales appeared as a triangular or irregular bone fragment of the distal acromion, and forms a pseudo-acromioclavicular joint with the acromion. Eleven cases were edematous os acromiale, 11 cases were displaced os acromiale. In the os arcomiale group, 17 had supraspinatus tear, 1 had supraspinatus tendinitis, 11 had infraspinatus tear, and 4 had infraspinatus tendinitis. In the no os arcomiale group, 11 had supraspinatus tear, 2 had supraspinatus tendinitis, 5 had infraspinatus tear, and 1 had infraspinatus tendinitis. No statistically significant difference between the os arcomiale group and no os arcomiale group regarding the rotator cuff tear, supraspinatus and infraspinatus injury (P>0.05). In the 11 cases of edematous os arcomiale, 10 had supraspinatus tear and 7 had infraspinatus tear. In the 10 cases of non-edematous os acromiale, 7 had supraspinatus tear and 4 had infraspinatus tear. No statistically significant difference was noted between the edematous os acromiale and non-edematous os acromiale in terms of supraspinatus and infraspinatus tear (P>0.05). In the 11 cases of displaced os acromiale, 11 had supraspinatus tear and 9 had infraspinatus tear. In the 10 cases of non-displaced os acromiale, 6 had supraspinatus tear and 2 had infraspinatus tear. In the no os arcomiale group, 11 had supraspinatus tear and 5 had infraspinatus tear. There was a statistically significant increases in the prevalence of supraspinatus and infraspinatus tear in the displaced os acromiale group compared with non-displaced os acromiale group, the displaced os acromiale group and no os arcomiale group(P<0.05). CONCLUSION Shoulder MRI can very well depict os acromiale and can reveal associated abnormalities such as adjacent bone marrow edema, displaced deformity, and rotator cuff tear, and it can be used to assess the stability of the os acromiale. The presence of os acromiale may not increase the risk of supraspinatus and infraspinatus tear significantly. However, the presence of displaced os acromiale is at greater risk of supraspinatus and infraspinatus tear.
Acromion/diagnostic imaging* ; Adult ; Aged ; Female ; Humans ; Magnetic Resonance Imaging ; Male ; Middle Aged ; Rotator Cuff ; Rotator Cuff Injuries/diagnostic imaging* ; Shoulder

OBJECTIVETo investigate the possible effects of apolipoprotein A1 gene (APOA1) rs670 and rs5069 polymorphisms on plasma lipid profiles in healthy adolescents with different body mass index (BMI).

METHODSTotally 723 adolescents were divided into four groups according to their BMI: group 1[BMI =(17.80 ± 0.75)kg/m2], group 2[BMI = (19.39 ± 0.32) kg/m²], group 3[BMI = (20.68 ± 0.43) kg/m²], and group 4[BMI=(23.40 ± 2.05) kg/m²]. Height, weight, waist circumference, hip circumference, blood pressure, heart rate, plasma lipids, and blood glucose were determined, BMI and waist to hip ratio (W/H ratio) were calculated,and genome DNA was extracted for analyzing the genotypes of the APOA1 rs670 and rs5069 polymorphisms by polymerase chain reaction-restriction fragment length polymorphism.

RESULTSNo significant differences in height, weight, BMI, waist circumference, hip circumference, W/H ratio, blood pressure, heart rate, plasma lipids, and blood glucose between APOA1 rs670 or rs5069 genotypes were observed among group 1, group 2, and group 3. In group 4, A carriers of the rs670 polymorphism had significantly higher systolic blood pressure (P=0.017) and blood glucose levels (P=0.009) than the adolescents with the GG genotype. T carriers of the rs5069 polymorphism had significantly higher height (P=0.013), weight (P=0.011), and hip circumference (P=0.026) than the adolescents with the CC genotype.

CONCLUSIONSIn healthy adolescents with higher BMI, APOA1 rs670 polymorphism is associated with systolic blood pressure and blood glucose levels. The elevation of systolic blood pressure and blood glucose levels in A carriers of APOA1 rs670 polymorphism may be favorably modulated by weight loss.

Adolescent ; Apolipoprotein A-I ; genetics ; Body Mass Index ; Female ; Humans ; Lipids ; blood ; Male ; Polymorphism, Genetic