Abstract A rapid gas chromatographic ( GC ) method was established for the determination of short-chain fatty acids ( SCFAs ) in human feces. Feces samples were directly extracted by 1% HCl-75% ethanol solution, and then centrifuged at high speed for GC analysis. The chromatographic separation was performed on a DB-FFAP capillary column ( 30 m í 0 . 25 mm í 0 . 25 μm ) with a temperature program ( initial temperature at 50℃ held for 1 min, ramped to 190℃ at 10℃/min ) . The injection port temperature was 250℃ with split ratio at 50:1 . The carrier gas was high purity nitrogen with a constant linear velocity at 1. 0 mL/min. A flame ionization detector was employed to quantify SCFAs. The proposed method had been certified by systematic method validation, and used to determine feces samples. Subsequently, the health volunteer and colorectal cancer patient groups could be distinguished successfully by multivariate statistical analysis. Compared with health volunteers, the acetic acid and butyric acid of feces from colorectal cancer patients were reduced obviously, indicating that SCFAs particularly butyric acid could be considered as candidate markers for colorectal cancer diagnosis. In summary, this study provides a rapid method for the determination of SCFAs in feces form health volunteers and colorectal cancer patients. The method had a prospect for screening and diagnosing colorectal cancer rapidly.

Objective To evaluate the role of via-anal ileus tube drainage in the therapy of acute low colorectal obstruction caused by cancer in elderly patients.Methods Totally 31 elderly patients with acute obstruction caused by colorectal cancer were treated by this modality.With the help of colonscopy and radiograph,the ileus tube was inserted into the proximal bowel of the obstructive side via anus.Then we irrigated and drain through the tube.At the same time we observed the patients' symptoms,abdominal girth,plain abdominal radiograph.After the relief of the acute obstruction,operations were processed.Results After 7-14 days treatment,the acute obstruction was relieved and emergency operation was avoided.The cure rate of 1-staged resection was 93.5％ (29/31),the rate of 1-staged anastomosis was 68.9％ (20/29),the rate of complication was 19.4％ (6/31),no anastomotic leakage and no perioperative death occurred.Conclusions The via-anal tube drainage in the treatment of the elderly patients with acute obstruction is safe,effective,and the trauma is small.It would be the first choice to relieve the acute obstruction in future.

Aim To screen a more suitable transfection recep-tor,and improve the efficiency of constructing cell lines highly expressing human peptide transporters 1 (hPepT1 ).Methods The recombinant plasmid pcDNA3.1 (+)-hPepT1 was transfect-ed into MDCK cells and HeLa cells by LipofectamineTM 2000 transfection reagent,respectively.The monoclonal cells were se-lected and cultured.Expression of hPepT1 mRNA and protein were determined by qRT-PCR and Western blot,respectively. The uptake capacity of Glysar in transfected cells was examined. Results Compared with wild type cells,the expression of hPepT1 and the uptake of Glysar in transfected MDCK cells and HeLa cells significantly increased (P <0.05).Although the up-take of Glysar in HeLa cells was higher than that of MDCK cells,on the contrary,the expression of hPepT1 and the uptake of Glysar in MDCK-hPepT1 cells was higher than that of HeLa-hPepT1 cells.Conclusion MDCK cells may serve as a more suitable transfected receptor for the construction of a cellular model with high expression of hPepT1 ,which would make the construction of a cell model highly expressing hPepT1 more effi-cient.

Objective To evaluate the safety,feasibility and value of treating familial adenomatous polyposis (FAP) with laparoscopic assisted total colorectal resection.Methods Thirty six patients with familial adenomatous polyposis from June 2009 to May 2014 were reviewed retrospectively.16 FAP patients underwent laparoscopic assisted total colorectal resection and 20 FAP underwent traditional laparotomy.The clinical data,including short term follow up outcomes,safety,and recovery were analyzed retrospectively.Results Laparoscopic assisted total colorectal resection was performed successfully on 16 patients without severe complications.The mean operation time of the laparoscopic group was (178 ± 33) min,the mean operative blood loss (72 ± 30)ml in the laparoseopic group were not significantly different when compared with the mean operation time (159 ± 24) min and the mean operative blood loss (110 ± 50) ml in the conventional group.Incision length (5.6 ± 1.1) cm,the intestinal function recovery time (2.7 ± 0.9) d,hospital stay after surgery (8.1 ± 1.6) d in the laparoseopic group were significantly different when compared with incision length (15.8 ± 1.8) cm,the intestinal recovery time (3.8 ±0.9) d,hospital stay after surgery (9.9 ± 1.6) d in the conventional group (P ＜ 0.05).There was no severe complications in neither group nor local recurrence,distant nletastases or death found during the follow-up of 6-56 months.Conclusions Laparoscopic assisted total colorectal resection for FAP can be performed safely and effectively with the advantages of minimal invasion,quick recovery and good short term effect.

Abstrate:Aim To construct HEK293 cell line with stable and high expression of sodium taurocholate cotransporting polypeptide (NTCP ) efficiently and rapidly.Method Vector expressing EGFP-NTCP fusion protein was constructed and verified by DNA sequencing.The pEGFP-NTCP expression vector was transfected into HEK293 cells by FuGENE 6 transfection reagent. The transfected cells with high expression of green fluorescent protein were selected using fluorescence microscope for screening of G418 for 14 days to obtain cell lines stably and highly expressing NTCP.NTCP expression was detected by RT-PCR,qRT-PCR, Western-blot and the uptake experiment of taurocholic acid.Re-sult RT-PCR,qRT-PCR,Western-blot and the uptake experi-ment revealed that compared to the control cells,the expression of NTCP was significantly positive (P<0.01)in stable trans-fected cells showing green fluorescence (P<0.05 ).Conclusion The HEK293 cell line with stable and high expression of NTCP has been established efficiently and rapidly,which provides a cellular model for the study of the mechanism of the uptake of bile acid derivatives.

Objective To investigate the modulation of EPCs by interleukin 1β (IL-1β) and p38 mitogen activated protein kinase (p38MAPK) and the pathogenesis resulting from their dysdifferenfiation after trauma.Method Thirty pigs were divided into a control group (n = 15) and a multiple organ dysfimction syndrome (MODS) group (n = 15), the latter of which were subjected to a "two-hit" injury including hemon'hagic shock and endotoxemia. Phosphorylation of p38MAPK in peripheral blood mononuclear cells was monitored by western blotting. The concentration of IL-1β in peripheral blood plasma was determined by ELISA and the numbers of EPCs with FCM in peripheral blood plasma were monitored. The morbidity rates in the two groups were compared by chi square test. The levels of phosphorylation of p38MAPK in peripheral blood mononuclear cells, the concentmtions of IL-1β in peripheral blood plasma and the numbers of EPCs in the peripheral blood were compared between groups using with Student's t lest. Results The level of p38MAPK phosphorylation was more augmented and the concen-tration of IL-1β higher in peripheral blood mononuelear cells and plasma from MODS pigs compared with those from control pigs; nevertheless the mauler of EPC conspicuously decreased in the peripheral blood (P <0.01). The morbidity rate in the MODS group was much higher than that in the control group (P < 0.01). There were fewer EPCs in the peripheral blood of animals in group M than in the peripheral blood of animals in group C (P <0.01). Conclusions p38MAPK phosphorylation is important for the pathogenesis of MODS. p38MAPK phospho-rylation might cause the concentration of IL-1β in the peripheral blood plasma to rise and could cause a drop in the numbers of EPCs, thereby aggravating the inflanmmatory reaction in MODS.

Objective:To investigate the effect of total anatomical reconstruction (TAR) during robot-assisted radical prostatectomy (RARP) .Methods:The clinical data of 99 patients with RARP performed by a single doctor in our hospital from January 2018 to January 2021 were analyzed retrospectively.There were 38 patients in the TAR+ vesicourethral anastomosis (VUA) group and 61 patients in the VUA group. There were no significant differences between the two groups in the age of patients [ 65.5 (60.8, 71.0) years vs. 66.0 (61.5, 69.0) years], body mass index[ (24.92±2.65) kg/m 2 vs. (25.51±2.80) kg/m 2], prostate volume [28.13 (25.21, 36.53) ml vs. 26.33 (19.75, 47.84) ml], PSA [15.67 (9.02, 31.49) ng/ml vs. 14.58 (9.23, 30.06) ng/ml], neoadjuvant therapy [50.0% (19/38) vs. 63.9% (39/61)], Gleason score (6/7/8/9-10 scores: 8/16/5/9 cases vs. 16/25/9/11 cases) and clinical T stage (T 1/T 2/T 3 stage: 4/29/5 cases vs. 3/53/5 cases)(all P>0.05). The TAR technique was performed as follows. ①The two layers of posterior reconstruction involved the residual Denonvilliers fascia, the striated sphincter and medial dorsal raphe (MDR), and the vesicoprostatic muscle (VPM), the fascia which was 1-2 cm from the cranial side of the bladder neck and MDR. ②The one layer of anterior reconstruction involved detrusor apron, tissues around the urethra and the visceral and parietal layers of the endoplevic fascia. The VUA technique was suturing the bladder neck and urethra consecutively. Perioperative indexes were compared between the two groups. Results:All 99 operations were successfully completed. There were no statistically significant differences between the TAR+ VUA and VUA groups in operation time [ (174.16±47.21) min vs. (188.70±45.39) min], blood loss [ 50 (50, 100) ml vs. 100 (50, 100) ml], incidence of postoperative complications [10.5% (4/38) vs. 14.8% (9/61)], phathological T stage [pT 2/pT 3~4 stage: 25/12 cases vs. 42/19 cases, P=0.895], and the time of indwelling catheter [ 21.0 (19.0, 21.0) d vs. 21.0 (21.0, 21.0) d] (all P>0.05). The difference in postoperative length of stay between the two groups was statistically significant[6.0 (5.0, 6.0) d vs. 7.0 (6.0, 7.5)d, P<0.001]. Follow-up was performed for 1 year after surgery. The recovery rate of urinary continence 3 months after surgery in TAR+ VUA and VUA groups were 86.8% (33/38) vs. 65.6% (40/61), which were statistically significant( P=0.019). There were no significant differences between TAR+ VUA and VUA groups in recovery rate of urinary continence 1 months after surgery [47.4% (18/38) vs. 45.9% (28/61)], 6 months after surgery [94.7% (36/38) vs. 85.2% (52/61)], and 12 months after surgery [94.7% (36/38) vs. 93.4% (57/61)] (all P>0.05). Conclusions:TAR technique has good surgical safety, and can promote recovery of early urinary continence after RARP.

The human Gadd45 protein family plays critical roles in DNA repair, negative growth control, genomic stability, cell cycle checkpoints and apoptosis. Here we report the crystal structure of human Gadd45γ [corrected], revealing a unique dimer formed via a bundle of four parallel helices, involving the most conserved residues among the Gadd45 isoforms. Mutational analysis of human Gadd45γ [corrected] identified a conserved, highly acidic patch in the central region of the dimer for interaction with the proliferating cell nuclear antigen (PCNA), p21 and cdc2, suggesting that the parallel dimer is the active form for the interaction. Cellular assays indicate that: (1) dimerization of Gadd45γ [corrected] is necessary for apoptosis as well as growth inhibition, and that cell growth inhibition is caused by both cell cycle arrest and apoptosis; (2) a conserved and highly acidic patch on the dimer surface, including the important residues Glu87 and Asp89, is a putative interface for binding proteins related to the cell cycle, DNA repair and apoptosis. These results reveal the mechanism of self-association by Gadd45 proteins and the importance of this self-association for their biological function.
Amino Acid Motifs ; Animals ; Apoptosis ; radiation effects ; CDC2 Protein Kinase ; Cell Cycle ; Cell Survival ; Crystallography, X-Ray ; Cyclin B ; metabolism ; Cyclin-Dependent Kinase Inhibitor p21 ; metabolism ; Cyclin-Dependent Kinases ; HeLa Cells ; Humans ; Intracellular Signaling Peptides and Proteins ; chemistry ; genetics ; metabolism ; Mice ; Mutagenesis, Site-Directed ; Mutation, Missense ; Proliferating Cell Nuclear Antigen ; metabolism ; Protein Binding ; Protein Interaction Domains and Motifs ; Protein Multimerization ; Protein Structure, Quaternary ; Ultraviolet Rays

Objective:To investigate the relationship between the positive surgical margin and clinical factors such as neoadjuvant hormonal therapy after robot-assisted laparoscopic radical prostatectomy (RARP) in high-risk patients with prostate cancer.Methods:The clinical data of 164 patients with high-risk prostate cancer being performed RARP by one surgeon were analyzed retrospectively in our hospital from January 2016 to January 2022. The mean patient’s age was (65.3±6.2) years old, mean body mass index (BMI) was (25.6±3.0) kg/m 2, the median value of total prostate specific antigen (tPSA) before operation was 18.6(11.3, 31.3)ng/ml, the median value of Gleason score before operation was 7 (7, 8), the median value of prostate volume was 29.3 (22.4, 40.2) ml, and the clinical stage was T 2aN 0M 0-T 4N 0M 0. 80 patients with prostate cancer were treated with neoadjuvant endocrine therapy. All of them were treated with complete androgen blockade with a median course of 3 months. Univariate analysis was used to analyze the correlation between age, BMI, prostate volume, neoadjuvant hormonal therapy, preoperative tPSA, clinical stage, Gleason score before operation and positive surgical margin. Then multivariate logistic regression was used to further analyze the independent risk factor of positive surgical margin after RARP. Results:The postoperative pathological diagnosis included pT 2 stage in 111 cases (67.7%), pT 3a stage in 15 cases (9.1%), pT 3b stage in 25 cases (15.2%), pT 4 stage in 13 cases (7.9%). No lymph node metastasis was noticed in all patients. The Gleason scores included 6 in 11 cases (6.7%), 3+ 4 in 26 cases (15.9%), 4+ 3 in 36 cases (22.0%), 8 in 17 cases (10.4%), 9-10 in 24 cases (14.6%), un-evaluation due to endocrine therapy in 50 (30.5%). The positive surgical margin of high-risk patients with prostate cancer was 44.5% (73/164). Univariate analysis showed that the neoadjuvant hormonal therapy, tPSA and clinical stage were correlated with positive surgical margin ( P<0.05). Multivariate logistic regression analysis showed that non-neoadjuvant hormonal therapy, preoperative tPSA>20ng/ml and clinical stage>T 2b were independent risk factors for positive surgical margin of high-risk patients with prostate cancer. Stratified analysis showed that when the preoperative tPSA was 10-20 ng/ml(21.1% vs.55.9%, P=0.014), the clinical stage was T 2c(29.6% vs.49.1%, P=0.040), the Gleason score before operation was 7(19.4% vs.54.1%, P=0.003), the positive surgical margin of high-risk patients in the neoadjuvant hormonal therapy group was significantly lower than that in the non-neoadjuvant hormonal therapy group ( P<0.05). Conclusions:Non-neoadjuvant hormonal therapy, preoperative tPSA>20 ng/ml and clinical stage>T 2b were independent risk factors for positive surgical margin of RARP in the high-risk patients with prostate cancer. For high-risk patients with preoperative tPSA of 10-20 ng/ml, clinical stage of T 2c and Gleason score before operation of 7, neoadjuvant hormonal therapy has important clinical significance in reducing the positive surgical margin of RARP.

In order to search for new adefovir analogues as anti-HBV agents with enhanced antiviral activity and hepatotrophic property,adefovir bis L-amino acid ester was used as lead compound to produce ten adefovir mono L-(thio)amino acid ester, mono bile acid ester derivatives(6a-6j). The design based on bile acid prodrug strategy,which can improve drug oral bioavaliability and liver-targeted enrichment by using enterohepatic circula-tion of bile acid.Sub-structure combination method was adopted to introduce L-(thio)amino acid ester and bile acid ester fragments on the phosphonate functionality of adefovir. The structures of target compounds were confirmed by 1H NMR, 13C NMR,ESI-MS and ESI-HRMS.HepG 2.2.15 cell were used for in vitro anti-HBV activity assessment.Compound 6c with high antiviral activity(EC500.92μmol/L,SI 512.63)was further investi-gated for its tissue distribution in mice.The results showed that content of compound 6c in liver was higher than that of adefovir dipivoxil,and in contrast its content in kidney was lower than that in positive control at all time points(0.25-12 h).Compound 6c exhibits higher antiviral activity,selective index and higher liver distribution,making it a potential anti HBV agent for further investigation.