Objective: This research explores the relationship between the immuno-suppression function of regulatory T cells (Treg) in the ascites of ovarian cancer (OC) patients, the clinico-pathologic features of these patients, and the correlation of the function of Treg with initial treatment and relapse status of the patients to further investigate the specific mechanism of immuno-regulatory func-tion of CD4+ CD25+ Treg in the ascites of OC. Methods: Immuno-magnetic activated cell sorting (MACS) was conducted to sort CD4+CD25+Treg and autologous CD4+CD25-Treg from the ascites of 28 OC patients. Carboxyfluorescein-diacetate succinimidyl ester (CFSE) was used to label the autologous CD4+CD25-Treg. These labeled cells were then used as controls and co-cultured with autologous CD4+CD25+Treg at the ratio of 1∶1 or 1∶2. The mean inhibition ratio of Treg in specimens to the proliferation of autolo-gous CD4+ CD25-Treg was calculated after the flow cytometry of the CFSE expression and Modfit software analysis of the CD4+CD25-Treg proliferation index (PI) were performed. Anti-IL-10 and/or anti-TGF-β1 antibodies were neutralized to investigate whether the CD4+CD25+Treg-mediated immuno-suppression escaped through the ascites can produce a marked effect by the inhibitory cyto-kine IL-10 or TGF-β1. Results: The mean inhibition ratio of CD4+ CD25- Treg in the ascites of stage Ⅲ to Ⅳ OC patients was (75.72±17.04)%, which is significantly higher than that of stageⅠtoⅡOC patients (59.61±16.97)%;P<0.05. In addition, Treg in the as-cites of OC patients with recurrent disease showed a significantly higher inhibition ratio than that of patients with primary disease;P<0.001. Moreover, Treg in groups added into neutralizing anti-IL-10 and/or anti-TGF-β1 antibodies displayed significantly lower depres-sant effect than the control group;P<0.05. Conclusion:The immuno-suppression of CD4+CD25+Treg in the ascites of OC patients is correlated with the tumor staging and status of the primary or recurrent diseases. Moreover, Treg may indicate a suppressor function by secreting cytokine IL-10 and TGF-β1.

Objective:CD4+CD25+regulatory T cells (Treg) may contribute to tumor progression by suppressing antitumor im-munity. The function of Treg in antitumor immunity regulation in the peritoneal microenvironment of ovarian cancer (OC) was investi-gated and compared with the circulating Treg to elucidate OC immune escape. Methods: Flow cytometry was used to determine the proportion of CD4+CD25+T cells in CD4+T cells in ascites of 27 patients with OC and in peripheral blood lymphocytes of 28 patients with OC. The samples were analyzed and classified in three stages:primary disease (PD), after chemotherapy (AC), and recurrence dis-ease (RD), according to the clinical conditions of the OC patients upon donating the samples. The percentage of Treg in the three groups was determined in ascites and blood. CD4+CD25+T cells were isolated from ascites and peripheral blood of patients with OC us-ing magnetic sorting (MACS) system. The cells were then tested for regulatory function through coculture with carboxyfluorescein diac-etate succinimidyl ester-labeled autologous CD4+ CD25- responder cells. Results:The proportion of CD4+ CD25+T cells in CD4+T cells significantly increased in ascites (28.25%± 14.06%) compared with that in blood (14.6%± 4.74%;P<0.0001). The Treg in ascites and blood in AC showed higher proportion (P<0.0001) than those in the PD and RD;the proportion in RD was higher than that in PD (P<0.0001). Moreover, the Treg in ascites mediated a significantly higher suppression compared with the Treg in peripheral blood (P<0.001). Conclusion:The frequency and suppressor function of Treg were significantly higher in ascites than in peripheral blood. This finding suggests more possibility for escape immune surveillance in the peritoneal microenvironment. Moreover, the proportion of Treg in AC was higher than that in PD or RD;the proportion in RD was higher than that in the PD. Chemotherapy may favor the expansion of Treg, which may promote the recurrence of cancer.

Objective:The objective of this research is to study the serum level of the high-mobility group protein B1 (HMGB1) in human ovarian tumor (OvCa) and in a healthy control. This study also aims to identify different HMGB1 levels before and after sur-gery and to explore the inhibitory effect of HMGB1 gene silencing in the proliferation and invasion ability of OvCa. Methods: En-zyme-linked immunosorbent assay was used to measure the serum level of HMGB1 in OvCa patients and healthy subjects. Lentivirus vector with HMGB1 shRNA was constructed and used to infect OvCa cells. The expressions of HMGB1 mRNA and protein were test-ed by real-time PCR and Western blot. Cell proliferation was detected using the Cell Counting Kit-8 assay, whereas cell invasion and migration were detected by Transwell assay. Results:The serum level of HMGB1 was more elevated in patients with malignant diseas-es compared with individuals with benign diseases and the control groups. In the malignant group, the serum level of HMGB1 de-creased noticeably after therapy. Down-regulation of HMGB1 expression resulted in the inhibition of the biological behavior and metas-tasis of ovarian cancer cells. Conclusion: HMGB1 is closely associated with clinicopathologic features of OvCa. Knockdown of HMGB1 expression can significantly inhibit cell proliferation, cell migration, and cell invasion of OvCa. These findings indicate that HMGB1 can function as a therapeutic target for ovarian neoplasm in the future.

Objective To observe the influence of low protein diet with α-keto acid on kidney sclerosis and renin-angiotensin system in renal ablation rats. Methods Chronic renal failure rat model was established by renal ablation in 30 male SD rats,then the animals were randomly assigned to the following diet groups:normal protein group (NPD:18%casein protein),low protein group (LPD:6%casein protein) and supplemented low protein group (LK:5%casein protein+1%α-keto acids).Ten male SD sham-operated rats received 18%casein protein as control.All the rats were killed at the end of the 12th week.Pathologic changes were assessed by PAS staining.Ang II in homogenate and plasma were measured by radioimmunoassay and ELISA respectively.Immunohistochemistry and Western blotting were used to detect the protein expression of TGF-β1,renin and AT1R.Real-time PCR was used to detect the gene expression of renin and ATla,the main subtype of AT1 receptor. Results Body weight,total protein and serum albumin had not significant difference among the four groups(all P>0.05).Serum creatinine and proteinuria of nephrectomized rats were significantly higher compared to the control group (all P<0.05).Proteinuria of the LK group was lower than that of NPD and LPD groups (all P<0.05).Pathological results indicated fibrosis indices were significantly improved after LPD and LK intervention.Expressions of renin,Ang II and AT1R in LK group were significantly lower than those in NPD group (all P<0.05). Conclusions Low protein diet with α-keto acids supplement therapy exhibits renal protective effects of reducing urine protein excretion and improving renal fibrosis,which might be related to the attenuation of local renin-angiotensin system in activity nephrectomized rats.

Objective To observe the alterations of Sirt1 and mTOR activity in gastrocnemius muscles of mice with sarcopenia. Methods Aged (26 months old) and young (6 months old) male C57BL/6J mice (n=16,respective-ly) were randomly arranged into 2 groups respectively, i. e. middle-aged control ( MC) , middle-aged suspension ( MS) , aged control( AC) and aged suspension( AS) . Aged mice were hindlimb suspended for two weeks to estab-lish a sarcopenic model. Kits and Western blot were used to detecte related factors and protein expression of Sirt1 and mTOR pathway. Results 1) the content of NAD+and the ratio of NAD+/NADH: AC group was significantly lower than MC group ( P＜0.01 or P＜0.05) , HLS group was significantly less than the control group ( P＜0.01 or P＜0.05) . 2) Sirt1 expression: AC group was significantly higher than MC group ( P＜0.05) , and HLS group was significantly higher than that of the control group (P＜0.05).3)Sirt1 downstream substrates (PGC-1α, FOXO1 and P53), NAMPT protein expression and Sirt1 activity: AC group was significantly lower than MC group ( P＜0.05 or P＜0.05 or P＜0.01) , HLS group was significantly less than the control group ( P＜0.05 or P＜0.05 or P＜0.05) . 4) phosphorylation of mTOR, p70S6K and 4E-BP1: AC group was significantly higher than MC group ( P＜0.05) , HLS group was significantly less than the control group ( P＜0.05) . Conclusions Sirt1 and mTOR are involved in the pathogenesis of mice with sarcopenia.

Fifteen cassaen-type diterpenes were isolated from the 95% ethanolic extract of the seeds of C. minax through various chromatographic techniques. Their structures were identified on the basis of spectroscopic data as pulcherralpin (1), caesalpinin ML (2), chamaetexane C (3), chamaetexane D (4), 6β, 18-diacetoxycassan-13, 15-diene (5), neocaesalpin K (6), neocaesalpin MP (7), neocaesalpin M (8), neocaesalpin Q (9), neocaesalpin P (10), neocaesalpin R (11), caesaldekarin D (12), caesaldekarin A (13), caesaldekarin b (14), 3β,6α-diacetoxyvouacapane (15). Among them, compounds 14, 9-11 were isolated from this plant for the first time.
Caesalpinia ; chemistry ; Diterpenes ; chemistry ; isolation & purification ; Drugs, Chinese Herbal ; chemistry ; isolation & purification ; Magnetic Resonance Spectroscopy ; Molecular Structure ; Seeds ; chemistry ; Spectrometry, Mass, Electrospray Ionization

OBJECTIVETo compare robot-assisted laparoscopic radical prostatectomy (RALRP) with laparoscopic radical prostatectomy (LRP) in the treatment of prostate cancer and investigate the clinical application value of RLRP.

METHODSWe retrospectively analyzed 70 cases of prostate cancer treated by RALRP and another 32 cases treated by LRP. We compared the operation time, intraoperative blood loss and transfusion, catheter-indwelling time, postoperative hospital stay, incisal margin positive rate, biochemical recurrence, and normal postoperative urinary continence and penile erectile function between the two groups of patients.

RESULTSAll the operations were successfully accomplished. RALRP exhibited a significant superiority over LRP in intraoperative blood loss and transfusion, catheter-indwelling time, and postoperative hospital stay, urinary continence and erectile function (P < 0.05).

CONCLUSIONRobot-assisted laparoscopic radical prostatectomy, with its advantages of few postoperative complications and well-preserved urinary continence and penile erectile function, is an effective, safe and minimally invasive surgical option for prostate cancer.

Blood Loss, Surgical ; Humans ; Laparoscopy ; Length of Stay ; Male ; Operative Time ; Penile Erection ; Postoperative Complications ; Postoperative Period ; Prostatectomy ; methods ; Prostatic Neoplasms ; surgery ; Retrospective Studies ; Robotic Surgical Procedures ; methods

Objective To observe the changes of renin-angiotensin system (RAS) in cultured mesangial cells by serum from 3/4 nephrectomized rats feeding with low protein diet and α-keto acid. Methods Thirty male SD rats received 3/4 nephrectomy (Nx) were placed on 18%normal protein diet (NPD,n=10),6% low protein diet(LPD,n=10) or 5% low protein plus 1%α-keto acid diet (LK,n=10) flor 12 weeks.Ten male SD sham-operated rats fed with 18% normal protein diet were used as control (sham group).In addition,mesangial cells were cultured in sera (10%) collected from above animals treated with or without losartan (0.02 mmol/L)for 48 hours.ELISA was applied to detect the level of Ang II,TGF-β1 and fibronectin (FN) in cell medium.Westem blotting was used to determine the protein level of ATI receptor (AT1R)and real-time PCR was used to detect the mRNA level of AT1R,TGF-β1 and FN. Results (1) Nutritional indices including body weight,total protein and albumin had no significant difference in each group. (2) Serum creatinine and 24 h pruteinuria were significantly inceased in nephrectomized groups compared to sham group(P<0.05,respectively).24 h proteinuria was greatly lower in LK group than that in NPD and LPD groups(P<0.05,respectively).(3)LK greatly decteased the level of Ang II[NPD(12.70±0.12)mg/g protein;sham(8.04±0.62)mg/g protein]in supernatant as well as the protein and mRNA expression of AT1R in cultured mesangial cells (P<0.05).(4)NPD serum directly induced higher secretion[FN:sham(20.58±0.46)g/g protein,NPD (39.84±0.06)g/g protein;TGF-β1:sham(10.12±O.56)mg/g protein,NPD(83.85±3.58)mg/g protein] and mRNA expression of FN and TGF-β1 compared with sham group (P<0.05).LPD decreased these increment (P<0.05) and LK showed stronger inhibitory effect (P<0.05). (5)Losartan application sharply reduced FN and TGF-β1 production both in supematant and in mRNA expression in NPD serum treated cells (P<0.05,respectively). Conclusion Low protein diet with α-keto acids supplement directly inhibits the RAS in mesangial cells which may contribute to its beneficial effect on the kidney.

OBJECTIVETo evaluate the application of traditional cytomorphology, telomerase activity analysis and immunocytochemistry in cytopathologic diagnosis of pleural effusion and bronchoalveolar lavage samples.

METHODSA total of 123 agar-paraffin double-embedded pleural effusion and bronchoalveolar lavage fluid samples were enrolled into study. The cytomorphologic features were reviewed and correlated with immunocytochemical findings and telomerase activity.

RESULTSTelomerase activity was detected in 53 specimens using the real-time telomeric repeat amplification protocol. Amongst the cases studied, 39 samples (31.7%) contained overtly malignant cells while 20 cases (16.0%) were equivocal by conventional cytology. After verification by immunocytochemistry and clinical follow-up data, the diagnostic accuracy of telomerase activity and cytology was 87.0% and 82.1%, respectively. The sensitivity (97.6%) and specificity (100.0%) of cytology examination, when combined with telomerase activity analysis, were greater than those of cytology examination or telomerase activity analysis alone.

CONCLUSIONSTelomerase activity analysis can be used as an adjunctive investigative tool in cytology assessment of pleural effusion and bronchoalveolar lavage samples. The diagnostic accuracy can be further improved with the application of immunocytochemistry on agar-paraffin double-embedded cell block tissues.

Breast Neoplasms ; diagnosis ; enzymology ; pathology ; Bronchoalveolar Lavage Fluid ; chemistry ; Female ; Follow-Up Studies ; Humans ; Immunohistochemistry ; Lung Neoplasms ; diagnosis ; enzymology ; pathology ; Pleural Effusion ; diagnosis ; enzymology ; pathology ; Pleural Effusion, Malignant ; diagnosis ; enzymology ; pathology ; Sensitivity and Specificity ; Telomerase ; metabolism

Objective - To investigate the effect of lung flora dysbiosis on the process of pulmonary fibrosis and lung epithelial （ ） Methods - mesenchymal transition EMT in mice with silicosis. Male C57BL/6 mice of specific pathogen free grade were ， ， ， （ ） randomly divided into the blank control group silicosis model group solvent control group vancomycin VM + ampicillin （ ） ， （ ） （ ） ， AMP group metronidazole MNZ + neomycin NEO group and mixed treatment group 12 mice in each group. Except for ， ， the blank control group which was given 20.0 µL of 0.9% NaCl solution the other five groups of mice were dosed with 20.0 µL of silica dust suspension at a mass concentration of 250.0 g/L using a single tracheal drip to establish the silicosis mouse model. ： The intranasal drip method was used to treat silicosis mice in each group as following mice in the solvent control group were - ； ； given double distilled water mice in the VM+AMP group were given VM at a mass concentration of 0.5 g/L and AMP at 1.0 g/L ； mice in the MNZ+NEO group were given MNZ at a mass concentration of 1.0 g/L and NEO at 1.0 g/L mice in the mixed ， treatment group were given the same doses of the four antibiotics mentioned above all in a drip volume of 50.0 µL. Silicosis ， ， mice were treated seven days and half an hour before silica dusting and 7 14 and 21 days after silica dusting. Mouse lungtissue was collected aseptically 28 days after silica dusting. Hematoxylin eosin and Masson trichrome staining methods were - used to observe the pathological changes. Western blotting was used to detect the relative protein expression of α smooth muscle （ - ）， - （ - ） （ ） actin α SMA E cadherin E CAD and vimentin VIM . Immunohistochemistry was used to detect the relative expression of - - E CAD and VIM. Real time fluorescence quantitative polymerase chain reaction was used to detect the expression levels of （Col1a2） Results collagen type Ⅰ alpha 2 mRNA in lung tissues. The histopathological results showed that the alveoli of the ， blank control group were thin and structurally intact with few surrounding infiltrating inflammatory cells and no abnormal ， distribution of collagen fibers. The alveoli of the silicosis model group were structurally disorganized with a large number of ， ， infiltrating inflammatory cells thickened alveolar walls and cellular fibrous nodules with abundant blue collagen deposit. In the ， ， VM+AMP group MNZ+NEO group and the mixed treatment group the inflammation and fibrosis were reduced with diferent degrees in the lung tissues compared to the silicosis model group and the solvent control group. The relative expression levels of - ， Col1a2 α SMA VIM protein and mRNA in lung tissues of mice in the silicosis model group were higher than those in the blank （ P ）， -CAD control group all <0.05 and the relative expression levels of E protein were lower than those in the blank control （P ） - ， Col1a2 group <0.05 . The relative expression levels of α SMA VIM protein and mRNA in lung tissues of mice in the MNZ+ （ P ）， -CAD NEO group and the mixed treatment group were lower all <0.05 and the relative expression levels of E protein were （P ）， Conclusion higher <0.05 when compared with the silicosis model group and the solvent control group. Pulmonary fibrosis ， - was reduced in silicosis mice with interventions in lung flora where anaerobic and gram negative bacteria affected pulmonary fibrosis and dysbiosis of the lung flora affected pulmonary EMT.