Piezo-Activated Atomic-Thin Molybdenum Disulfide/MXene Nanoenzyme for Integrated and Efficient Tumor Therapy via Ultrasound-Triggered Schottky Electric Field.

Monolayer molybdenum disulfide (MoS2 ) nanoenzymes exhibit a piezoelectric polarization, which generates reactive oxygen species to inactivate tumors under ultrasonic strain. However, its therapeutic efficiency is far away from satisfactory, due to stackable MoS2 , quenching of piezo-generated charges, and monotherapy. Herein, chitosan-exfoliated monolayer MoS2 (Ch-MS) is composited with atomic-thin MXene, Ti3 C2 (TC), to self-assemble a multimodal nanoplatform, Ti3 C2 -Chitosan-MoS2 (TC@Ch-MS), for tumor inactivation. TC@Ch-MS not only inherits piezoelectricity from monolayer MoS2 , but also maintains remarkable stability. Intrinsic metallic MXene combines with MoS2 to construct an interfacial Schottky heterojunction, facilitating the separation of electron-hole pairs and endowing TC@Ch-MS increase-sensitivity magnetic resonance imaging responding. Schottky interface also leads to peroxidase mimetics with excellent catalytic performance toward H2 O2 in the tumor microenvironment under mechanical vibration. TC@Ch-MS possesses the superior photothermal conversion efficiency than pristine TC under near-infrared ray illumination, attributed to its enhanced interlaminar conductivity. Meanwhile, TC@Ch-MS realizes optimized efficiency on tumor apoptosis with immunotherapy. Therefore, TC@Ch-MS achieves an integrated diagnosis and multimodal treatment nanoplatform, whereas the toxicity to normal tissue cells is negligible. This work may shed fresh light on optimizing the piezoelectric materials in biological applications, and also give prominence to the significance of intrinsic metallicity in MXene.

[Cognition and reflection on the "lateral ligament of rectum"].

As total mesorectal excision (TME) for rectal cancer is widely carried out in China, lateral ligament of rectum, as an important anatomical structure of the lateral rectum with certain anatomical value and clinical significance, has been the focus of attention. In this paper, by comparing and analyzing the characteristics about ligaments of the abdomen and pelvis, reviewing the membrane anatomy and the theory of primitive gut rotation, and combining clinical observations and histological studies, the author came to a conclusion that lateral ligament of rectum does not exist, but is only a relatively dense space on the rectal side accompanied by numerous tiny nerve plexuses and small blood vessels penetrating through it.

[Progress in clinical diagnosis and treatment of gallbladder cancer].

Due to the lack of effective early diagnosis and treatment, gallbladder cancer(GBC) remains a malignant tumor with extremely high malignancy and poor prognosis. Therefore, high quality studies are required to break through the bottleneck in GBC diagnosis and treatment. This article reviewed the domestic and foreign GBC research published in 2021, presenting a comprehensive summary of the important advances in the field of clinical diagnosis and treatment. Latest epidemiological data and risk factors, emerging diagnostic methods of peripheral blood laboratory tests and imaging, new pathologic classification system, hot topics and controversies of surgical treatment as well as the dynamics of systemic treatment of GBC are reviewed in the article. The present findings may contribute to a more efficient means of diagnosis and treatment for GBC and hold the promise of improved outcomes for patients with GBC.

Niobium carbide (MXene) reduces UHMWPE particle-induced osteolysis.

Joint replacement surgery is one of the orthopedic surgeries with high successful rates; however, wear debris generated from prostheses can ultimately lead to periprosthetic osteolysis and failure of the implant. The implant-derived particulate debris such as ultrahigh molecular weight polyethylene (UHMWPE) can initiate the local immune response and recruit monocytic cells to phagocytose particles for generating reactive oxygen species (ROS). ROS induces osteoclastogenesis and macrophages to secrete cytokines which ultimately promote the development of osteolysis. In this work, we develop the few-layered Nb2C (FNC) as an antioxidant which possesses the feature of decreasing the production of cytokines and inhibiting osteoclastogenesis by its ROS adsorption. Moreover, local injection of FNC attenuates the UHMWPE-induced osteolysis in a mouse calvarial model. In sum, our results suggest that FNC can be used for treating osteolytic bone disease caused by excessive osteoclastogenesis.

[Pathogens' distribution and changes of antimicrobial resistance in the bile of acute biliary tract infection patients].

Objective: To investigate the pathogens' distribution and antimicrobial resistance in the bile of acute biliary tract infection patients. Methods: The data of bile bacterial culture and drug sensitivity test of 223 acute biliary tract infection patients who underwent gallbladder puncture or endoscopic retrograde cholangiopancreatography drainage from July 2009 to July 2019 were analyzed retrospectively at Department of General Surgery,Xinhua Hospital,Affiliated to Shanghai Jiao Tong University School of Medicine.There were 141 males and 82 females with age of 67.3 years(range:28 to 93 years).Three to five milliliter of bile was extracted from each patient and sent to the laboratory for bacterial culture,identification and drug sensitivity test.The patients were divided into two groups according to the visiting time: the former group (n=124) was admitted from July 2009 to July 2014,and the latter group(n=99) was admitted from August 2014 to July 2019.The distribution of pathogenic bacteria and the changing trend of drug resistance rate of common bacteria in the two groups were compared.The results of drug sensitivity test were analyzed by WHONET software provided by WHO bacterial surveillance network.The drug resistance rates in different time periods were compared by χ2 test. Results: In this study,there were 147 cases of acute cholangitis and 76 cases of acute cholecystitis.A total of 376 strains of pathogenic bacteria were cultured.Among them,98 strains(26.1%) were gram-positive bacteria,269 strains(71.5%) were gram-negative bacteria and 9 strains(2.4%) were fungi.The top three gram-positive bacteria were Enterococcus faecium (49.0%,48/98),Enterococcus faecalis(20.4%,20/98),and Enterococcus luteus(7.1%,7/98).The top 5 gram-negative bacteria were Escherichia coli(33.5%,90/269),Klebsiella pneumoniae(13.8%,37/269),Pseudomonas aeruginosa(13.0%,35/269),Acinetobacter baumannii (12.6%,34/269),and Enterobacter cloacae(4.8%,13/269).From 2009 to 2019,there was no significant change in the proportion of gram-positive bacteria (former group vs. latter group: 25.3% vs. 28.2%) and gram-negative bacteria(former group vs.latter group: 74.7% vs. 71.8%) in the bile of patients with acute biliary tract infection.Gram-positive bacteria were mainly Enterococci(85.7%,84/98) and gram-negative bacteria were Escherichia coli(33.5%,90/269).Acinetobacter baumannii accounted for 7.8%(11/142) of gram-negative bacteria in the former group and 18.1%(23/127) in the latter group,an increase of 10.3% over previous five years.Pseudomonas aeruginosa had a downward trend,16.9% in the former group(24/142) and 8.7% in the latter group (11/127),the proportion decreased by 8.2%,and the other changes were not significant.The drug resistance rates of common gram-positive bacteria were relatively stable,and the drug resistance rates of Enterococcus faecium to many antibiotics were higher than those of Enterococcus faecalis.The resistance rates of gram-negative bacteria to most antibiotics showed an upward trend,among which Klebsiella pneumoniae showed an upward trend to most of antibiotics(former group: 0/15－4/13, latter group: 55.0%－70.0%; χ2=3.996－16.942, P=0.000－0.046).The drug resistance rates of Acinetobacter baumannii was generally higher,but there were no significant changes in the drug resistance rates of Acinetobacter baumannii between the two groups.The drug resistance rates of Pseudomonas aeruginosa to most antibiotics increased,and the overall drug resistance rates of Escherichia coli were stable and showed a slight upward trend. Conclusions: The main pathogens in bile of patients with acute biliary tract infection are gram-negative bacteria.The constituent ratio of various gram-negative bacteria had no significant change from 2009 to 2019,but the drug resistance rates shows an upward trend.Among the gram-negative bacteria, Escherichia coli is the most important pathogen,and the proportion has no significant change.The proportion of Acinetobacter baumannii in the former group was significantly higher than that in the former group.And the proportion of Pseudomonas aeruginosa has a decreased trend.

[Analysis of treatment modalities and prognosis of patients with gallbladder cancer in China from 2010 to 2017].

Objective: To evaluate the clinical characteristics and prognosis of gallbladder cancer (GBC) patients in China. Methods: This retrospective multicenter cohort study enrolled 3 528 consecutive GBC patients diagnosed between January 2010 to December 2017 in 15 hospitals from 10 provinces. There were 1 345 (38.12%) males and 2 183 (61.88%) females.The age of diagnosis was (63.7±10.8) years old (range: 26 to 99 years old) .There were 213 patients (6.04%) in stage 0 to Ⅰ, whereas 1 059 (30.02%) in stage Ⅱ to Ⅲ, 1 874 (53.12%) in stage Ⅳ, and 382 (10.83%) unavailable. Surgery was performed on 2 255 patients (63.92%) . Three hundred and thirty-six patients received chemotherapy or radiotherapy (9.52%; of which 172 were palliative); 1 101 (31.21%) received only supportive treatment.The patient source, treatment and surgery, pathology, concomitant gallstone, and prognosis were analyzed. Results: Among the 3 528 GBC patients, 959 (27.18%) were from East China, 603 (17.09%) from East-North China, 1 533 (43.45%) from Central China, and 433(12.27%) from West China. Among the 1 578 resectable tumor, 665 (42.14%) underwent radical surgery, 913 (57.86%) underwent surgery that failed to follow the guidelines.Eight hundred and ninety-one (56.46%) patients were diagnosed before surgery, 254 (16.10%) during surgery, and 381 (24.14%) after surgery (time point of diagnosis couldn't be determined in 52 patients) .Among the 1 578 patients with resectable tumor, 759 (48.10%) had concomitant gallstone.Among the 665 patients underwent radical surgery, 69 (10.4%) showed positive resection margin, 510 (76.7%) showed negative resection margin, and 86 (12.9%) unreported margin status.The 5-year overall survival rate (5yOS) for the 3 528-patient cohort was 23.0%.The 5yOS for patients with resectable tumor was 39.6%, for patients with stage ⅣB tumor without surgery was 5.4%, and for patients with stage ⅣB tumor underwent palliative surgery was 4.7%. Conclusions: More than half GBC patients in China are diagnosed in stage Ⅳ.Curative intent surgery is valuable in improving prognosis of resectable GBC.The treatment of GBC needs further standardization.Effective comprehensive treatment for GBC is in urgent need.

Circ_001680 aggravates the malignant process of gastric carcinoma by targeting MAP2.

OBJECTIVE: This study aims to explore the expression pattern and clinical significance of circ_001680 in gastric carcinoma (GC) process. PATIENTS AND METHODS: Circ_001680 levels in 40 pairs of GC and paracancerous ones were detected by quantitative real-time polymerase chain reaction (qRT-PCR). The relationship between circ_001680 and GC clinicopathological parameters was analyzed. AGS and SGC-7901 cells were used for constructing circ_001680 knockdown models by shRNA transfection. Proliferative and metastatic abilities in GC cells with circ_001680 knockdown were examined by cell counting kit-8 (CCK-8) and transwell assay, respectively. Dual-Luciferase reporter assay was conducted to clarify the interaction between circ_001680 and MAP2. Their co-regulation on GC process was detected through rescue experiments. RESULTS: Circ_001680 was highly expressed in GC tissues and cell lines. High level of circ_001680 predicted high incidences of lymphatic and distant metastasis, and poor prognosis in GC patients. Knockdown of circ_001680 suppressed proliferative and metastatic abilities in AGS and SGC-7901 cells. MAP2 was the target gene binding circ_001680, which was lowly expressed in GC. In addition, MAP2 was negatively correlated to circ_001680. Knockdown of MAP2 could abolish the suppressed proliferative and metastatic abilities in GC cells with circ_001680 knockdown. CONCLUSIONS: Circ_001680 is highly expressed in GC tissues and closely related to metastasis and prognosis in GC patients, which promotes the proliferative and metastatic abilities in GC cells by negatively interacting with MAP2.

[Laparoscopic gastroduodenostomy with manual suture for Billroth â anastomosis: a report of 36 cases].

Objective: To examine the clinical efficiency of laparoscopic gastroduodenostomy with BillrothⅠanastomosis with manual suture. Methods: The clinic data of 36 patients with gastric cancer who underwent laparoscopic gastroduodenostomy with Billroth Ⅰ anastomosis from November 2017 to September 2019 in Department of Gastrointestinal and Pancreatic Surgery, Zhejiang Provincial People's Hospital were analyzed retrospectively.There were 22 males and 14 females, aged (64.3±9.3) years(range: 43 to 80 years), underwent complete laparoscopic gastroduodenostomy. The laparoscopic manual suture was used for Billroth Ⅰ anastomosis. Results: All the laparoscopic radical gastrectomy and manual suturing gastroduodenostomy were successfully performed. The operation time was (226.7±40.4) minutes (range: 180 to 320 minutes), including (24.8±7.1) minutes (range: 15 to 48 minutes) for gastroduodenostomy.There was (3.8±0.9) days (range: 2 to 6 days) for anal exhaust, (5.7±2.0) days (range: 3 to 13 days) for extubation of gastric tube, and (10.3±3.1) days (range: 7 to 19 days) for hospitalization. There was no death in perioperative period. Postoperative pathological report showed 3 cases of highly differentiated adenocarcinoma, 5 cases of moderately differentiated adenocarcinoma, 22 cases of poorly differentiated adenocarcinoma and 6 cases of signet ring cell carcinoma, including 27 cases in T1 stage and 9 cases in T2 stage. The number of lymph nodes harvested was 36.4±8.9 (range: 23 to 60). Lymph node metastasis was positive in 7 cases and negative in 29 cases. TNM stage included 24 cases in ⅠA stage, 8 cases in ⅠB stage and 4 cases in Ⅱ stage. After the operation, the upper digestive tract radiography showed that the anastomosis opening was unobstructed without complications such as anastomotic stenosis. Conclusion: Laparoscopic gastroduodenostomy with Billroth Ⅰ anastomosis with manual suture is safe and feasible, has a good short-term effect.

[Diagnosis and treatment for unexpected gallbladder carcinoma(a retrospective study of 45 cases)].

Objective: To investigate the rationale for appropriate diagnostic methods and treatment protocols for unexpected gallbladder carcinoma(UGC). Methods: The clinical and pathological data of 45 patients with UGC admitted at Department of General Surgery, Xin Hua Hospital, Shanghai Jiao Tong University School of Medicine,from January 2008 to December 2017 were retrospectively collected and analyzed.There were 11 males(28.9%) and 34 females(71.1%),aged 68 years(range:27 to 68 years).And there were 20 cases who aged above 70 years. Twenty-four cases were diagnosed preoperatively as cholecystolithiasis plus chronic cholecystitis.Ten cases were diagnosed preoperatively as cholecystolithiasis plus actue cholecystitis.Six cases were diagnosed preoperatively as cholecystolithiasis plus choledocholith.Six cases were admitted because of gallbladder polyp and 1 case was admitted because of gallbladder adenomyomatosis. Results: Thirty-four patients with UGC received radical surgery.Among them,11 patients experienced postoperative complication and no posterative mortality occoured during hospital stay.Thirteen patients were diagnosed with T1b UGC, the harvested lymph node of Nx, N0, N1 and N2 was 2, 9, 1 and 1, respectively.In addition, 2 cases were identified to have local-regional tumor recurrence during our rescue radical surgery.The median overall survival time of the patients who did not receive radical surgery was 7 months(range:2-56 months).Nevertheless,the median overall survival time for patients diagnosed with T1, T2 and T3 tumors who received radical surgery, was 41 months(range: 19-82 months), 33.5 months(range: 31-36 months) and 17 months(range: 7-46 months), respectively. Conclusions: For patients with UGC, rescue radical surgery can achieve a better survival time.Furhtermore, our experience proved that rescue radical surgery for UGC is safe and feasible.Therefore,rescue radical surgery should be performed in patients with diagnose with UGC especially those T1b patients.

[The role of anatomical hepatectomy in the treatment of intrahepatic cholangiocarcinoma].

Objective: To evaluate the role of anatomical hepatectomy in the treatment of intrahepatic cholangiocarcinoma. Methods: The cases of intrahepatic cholangiocarcinoma who received curative surgery in two hospitals from 2010 to 2015 were analyzed retrospectively. Among the 98 patients enrolled in this study, 55 were male and 43 were female. The median age was 61 years. According to receiving anatomical hepatectomy or not, the 98 cases were divided into two groups: non-anatomical hepatectomy(n=30) and anatomical hepatectomy(n=68). The surgical results were compared between the two groups.Survival curves were plotted by the Kaplan-Meier method and compared by the log-rank test. The influence of each prognostic factor identified by univariate analysis was multivariate analysis by Cox's proportional hazard regression. Results: The duration of surgery was significantly prolonged in the anatomical hepatectomy group((196.4±94.9)minutes vs. (166.2±65.7)minutes, P=0.027), while there was no significant difference in terms of other surgical results such as intraoperative blood transfusion, postoperative morbidity and mortality rate. Compared to non-anatomical hepatectomy, anatomical hepatectomy significantly improved long-term survival results(14 months vs. 11 months)(χ2=4.641, P=0.031). Single variable analysis indicated that tumor differentiation, tumor numbers, T stage, N stage, anatomical hepatectomy and adjuvant therapy significantly affected overall survival. Multivariate analysis demonstrated that tumor numbers(HR=0.522, 95% CI: 0.259-0.974, P=0.042) and anatomical hepatectomy(HR=1.858, 95%CI: 1.092-3.161, P=0.022) were two independent prognostic factors for overall survival. Conclusion: Compared to non-anatomical hepatectomy, anatomical hepatectomy performed for intrahepatic cholangiocarcinoma is not only safe but also beneficial for long-term survival.

[Method and significance of specimens standardized pathological treatment in pancreaticoduodenectomy for pancreatic head cancer].

Pancreatic ductal adenocarcinoma is a highly aggressive disease with a grim prognosis. Surgical resection offers the best chance for long-term survival. Negative-margin resection still remains the goal, the influence of margin status on outcomes in pancreatic head carcinoma remains controversial, as conflicting data have been plagued by a lack of standardization in R0 resection and margin definitions, pathologic analysis, and reporting. In contrast to common belief, a high rate of R1 resections in pancreatic cancer is not a marker of low-quality surgery but rather of high-quality pathology. The international pathological consensus of pancreatic head carcinoma is still needed to fully understand the prognostic value of margin status in order to optimize treatment strategy for this disease.

[Effects of hypertonic sodium saline resuscitation on the liver damage of rats at early stage of severe scald].

Objective: To explore the effects of hypertonic sodium saline (HSS) resuscitation on the liver damage of rats at early stage of severe scald. Methods: Fifty-six SD rats were divided into sham injury group (SI, n=8), lactated Ringer's solution group (LRS, n=24), and group HSS (n=24) according to the random number table. Rats in group SI were sham injured without resuscitation, while rats in the other two groups were reproduced deep partial-thickness to full-thickness scald model with 30% total body surface area on the back. Rats in group LRS were resuscitated with LRS, while rats in group HSS were resuscitated with 300 mmol/L sodium ion solution according to the Parkland formula. Blood of abdominal aorta and liver of 8 rats in group SI immediately post injury and in the other two groups at post injury hour (PIH) 2, 8, and 24 respectively were collected. Then liver water content was determined by dry-wet weight method. Serum content of alanine aminotransferase (ALT) and aspartate transaminase (AST) was detected by automatic biochemical analyzer. Serum content of tumor necrosis factor α (TNF-α), interleukin-1 (IL-1), and high mobility group box 1 (HMGB1) was determined by enzyme-linked immunosorbent assay. Liver content of malondialdehyde (MDA) and superoxide dismutase (SOD) was detected by ultraviolet spectrophotometer. Pathologic changes of liver were observed by HE staining. Data were processed with one-way analysis of variance and SNK test. Results: (1) At PIH 2, 8, and 24, liver water content of rats in group LRS was higher than that in group SI and group HSS (P<0.05 or P<0.01). (2) At PIH 2, serum ALT content of rats in the three groups was similar (with P values above 0.05). At PIH 8 and 24, serum ALT content of rats in group HSS and group LRS was higher than that in group SI (P<0.05 or P<0.01), and serum ALT content of rats in group HSS was lower than that in group LRS (with P values below 0.01). At PIH 2, 8, and 24, serum AST content of rats in group HSS and group LRS was higher than that in group SI (with P values below 0.01). At PIH 2 and 8, serum AST content of rats in group HSS was lower than that in group LRS (P<0.05 or P<0.01). (3) At PIH 2 and 8, serum TNF-α content of rats in group LRS was (123±39) and (153±38) pg/mL respectively, higher than that in group SI [(60±18) pg/mL] and group HSS [(85±10) and (94±16) pg/mL respectively, with P values below 0.01]. At PIH 8, serum TNF-α content of rats in group HSS was higher than that in group SI (P<0.05). At PIH 24, serum TNF-α content of rats in the three groups was similar (with P values above 0.05). At PIH 2, 8, and 24, serum IL-1 content of rats in group LRS was (122±35), (141±30), and (122±31) pg/mL respectively, and that in group HSS was (80±12), (93±15), and (80±11) pg/mL respectively, all higher than that in group SI [(40±17) pg/mL, with P values below 0.01]; serum IL-1 content of rats in group HSS was lower than that in group LRS (with P values below 0.01). At PIH 2, serum HMGB1 content of rats in the three groups was similar (with P values above 0.05). At PIH 8 and 24, serum HMGB1 content of rats in group LRS was (0.386±0.146) and (0.590±0.188) ng/mL respectively, higher than that in group SI [(0.050±0.027) ng/mL] and group HSS [(0.143±0.038) and (0.309±0.095) ng/mL respectively, with P values below 0.01]. At PIH 24, serum HMGB1 content of rats in group HSS was higher than that in group SI (P<0.01). (4) At PIH 2, 8, and 24, liver MDA content of rats in group HSS and group LRS was higher than that in group SI and their liver SOD content was lower than that in group SI (with P values below 0.01); liver MDA content of rats in group HSS was lower than that in group LRS and their liver SOD content was higher than that in group LRS (with P values below 0.01). (5) Compared with those of rats in group SI, liver cells of rats in group LRS showed massive steatosis at each time point, and liver cell-edema appeared at PIH 8 and 24; while liver cells of rats in group HSS showed little steatosis only at PIH 8 and 24, and the liver cell-edema never appeared. Conclusions: Compared with LRS, HSS resuscitation can alleviate liver injury of rats at the early stage of severe scald through relieving inflammatory mediators and reducing degree of oxidative stress, etc.

FERMT1 mediates epithelial-mesenchymal transition to promote colon cancer metastasis via modulation of ß-catenin transcriptional activity.

We previously demonstrated that fermitin family member 1 (FERMT1) was significantly overexpressed in colon cancer (CC) and associated with poor metastasis-free survival. This study aimed to investigate the precise role of FERMT1 in CC metastasis and the mechanism by which FERMT1 is involved in the epithelial-mesenchymal transition (EMT). Correlations between FERMT1 and EMT markers (E-cadherin, Slug, N-cadherin and ß-catenin) were examined via immunohistochemistry in a cohort of CC tissues and adjacent normal colon mucosae. A series of in vitro and in vivo assays were performed to elucidate the function of FERMT1 in CC metastasis and underlying mechanisms. The upregulated expression of FERMT1 in CC tissues correlated positively with that of Slug, N-cadherin and ß-catenin, but correlated inversely with E-cadherin expression. Altered FERMT1 expression led to marked changes in the proliferation, migration, invasion and EMT markers of CC cells both in vitro and in vivo. Investigations of underlying mechanisms found that FERMT1 interacted directly with ß-catenin and activated the Wnt/ß-catenin signaling pathway by decreasing the phosphorylation level of ß-catenin, enhancing ß-catenin nuclear translocation and increasing the transcriptional activity of ß-catenin/TCF/LEF. Activation of the Wnt/ß-catenin pathway by CHIR99021 reversed the effect of FERMT1 knockdown, whereas inhibition of the Wnt/ß-catenin pathway by XAV939 impaired the effect of FERMT1 overexpression on EMT and cell motility. In conclusion, findings of this study suggest that FERMT1 activates the ß-catenin transcriptional activity to promote EMT in CC metastasis.

Leucine-rich repeat-containing G-protein-coupled receptor 5 is associated with invasion, metastasis, and could be a potential therapeutic target in human gastric cancer.

BACKGROUND: Leucine-rich repeat-containing G-protein-coupled receptor 5 (Lgr5), which is identified as a novel intestinal stem cell marker, is overexpressed in various tumours. In this study, we explore Lgr5 expression in gastric carcinoma and analyse its role in invasion, metastasis, and prognosis in carcinoma. METHODS: A combination of immunohistochemistry, western blotting, and quantitative reverse transcription-polymerase chain reaction were used to detect mRNA and protein expression levels of Lgr5 and matrix metalloproteinase 2 (MMP2). Small interfering RNA against Lgr5 was designed, synthesised, and transfected into AGS cells. The effects of Lgr5 siRNA on cell invasion were detected by transwell invasion chamber assay and wound healing assay. RESULTS: Leucine-rich repeat-containing G-protein-coupled receptor 5 expression was significantly higher in gastric carcinomas than in normal mucosa. Leucine-rich repeat-containing G-protein-coupled receptor 5 expression positively correlated with the depth of invasion, lymph node metastasis, distance of metastasis, and MMP2 expression levels. Multivariate analysis showed that Lgr5 had an independent effect on survival, and that it positively correlated with MMP2. Leucine-rich repeat-containing G-protein-coupled receptor 5 siRNAs inhibited Lgr5 mRNA and protein expression. Transwell assays indicated that these siRNAs resulted in significantly fewer cells migrating through the polycarbonate membrane, and wound healing assay also indicated that siRNAs decreased the migration of cells. Inhibition of Lgr5 resulted in a significant decrease in MMP2 and ß-catenin levels compared with those in controls. CONCLUSIONS: Leucine-rich repeat-containing G-protein-coupled receptor 5 was correlated with invasion and metastasis. Leucine-rich repeat-containing G-protein-coupled receptor 5 inhibition could serve as a novel therapeutic approach.

Ethyl pyruvate administration suppresses growth and invasion of gallbladder cancer cells via downregulation of HMGB1-RAGE axis.

High mobility group box B1 (HMGB1)-receptor for advanced glycation end products (RAGE) axis has been previously known to be involved in carcinogenesis and development of multiple malignancies. Some studies have confirmed that Ethyl pyruvate (EP), a potent inhibitor of HMGB1, exerts the therapeutic effects on metastatic live tumor from gastric cancer. However, the effects and possible molecular mechanisms of EP on gallbladder cancer (GBC) need to be further explored. In the present study, human GBC cell lines (GBC-SD and SGC-996) were treated with different concentrations of EP. Then, the expression levels of HMGB1, RAGE and some transcription factors were identified by Real-time PCR and Western blot assays. Cell proliferative activities indicated by MTT assay, invasive potential by Transwell assay and cell apoptosis and cycle distribution were performed for functional analysis of GBC cell lines in vitro. As a result, EP decreased the expression of HMGB11, RAGE, PCNA and matrix metallopeptidase-9 (MMP-9), while it increased the expression of p53. Moreover, EP administration decreased GBC cell proliferation, inhibited the invasive potential, and induced apoptosis and cycle arrest in S phase in GBC cells. In conclusion, EP administration inhibits growth and invasion of gallbladder cancer cells possibly via down-regulation of the HMGB1-RAGE axis, suggesting that EP may play a critical role in the treatment of cancer in conjunction with other therapeutic agents.

Side chain orientation in the selectivity filter of a voltage-gated Ca2+ channel.

Four glutamate residues (EEEE locus) are essential for ion selectivity in voltage-gated Ca(2+) channels, with ion-specific differences in binding to the locus providing the basis of selectivity. Whether side chain carboxylates or alternatively main chain carbonyls of these glutamates project into the pore to form the ion-binding locus has been uncertain. We have addressed this question by examining effects of sulfhydryl-modifying agents (methanethiosulfonates) on 20 cysteine-substituted mutant forms of an L-type Ca(2+) channel. Sulfhydryl modifiers partially blocked whole oocyte Ba(2+) currents carried by wild type channels, but this block was largely reversed with washout. In contrast, each of the four EEEE locus glutamate --> cysteine mutants (0 position) was persistently blocked by sulfhydryl modifiers, indicating covalent attachment of a modifying group to the side chain of the substituted cysteine. Cysteine substitutions at positions immediately adjacent to the EEEE locus glutamates (+/-1 positions) were also generally susceptible to sulfhydryl modification. Sulfhydryl modifiers had lesser effects on channels substituted one position further from the EEEE locus (+/-2 positions). These results indicate that the carboxylate-bearing side chains of the EEEE locus glutamates and their immediate neighbors project into the water-filled lumen of the pore to form an ion-binding locus. Thus the structure of the Ca(2+) channel selectivity filter differs substantially from that of ancestral K(+) channels.

A novel biodegradable system based on gelatin nanoparticles and poly(lactic-co-glycolic acid) microspheres for protein and peptide drug delivery.

Gelatin nanoparticle-poly(lactic-co-glycolic acid) (PLGA) microsphere composites were prepared by encapsulating protein-loaded gelatin nanoparticles in PLGA microspheres. This encapsulation was conducted by using a phase separation method and a solvent extraction method. The average diameter of the gelatin nanoparticle-PLGA microsphere composites is between 160 and 175 microm. Protein loading efficiency is 93.2% for the nanoparticle-microsphere composite prepared by the phase separation method, while it is 31.31% for the composite prepared by the solvent extraction method. Protein release experiments indicate that this new composite system possesses sustained release characteristics. This system also demonstrates the capability of preventing the denaturation of protein drugs.

Preparation and characterization of agarose hydrogel nanoparticles for protein and peptide drug delivery.

The purpose of this work was to develop and characterize a protein and peptide injectable drug delivery system in agarose hydrogel nanoparticles. The nanoparticles were prepared by using a new emulsion-converted-to-suspension in situ method. This is an emulsifier-free method that has advantages for protein and peptide drug encapsulations. Ovalbumin, used as a model protein drug, was successfully encapsulated into nearly spherical agarose hydrogel nanoparticles under mild conditions. The nanoparticles possessed a log-normal size distribution with an average size of 504 nm. They imbibed a large amount of water (66.85% to 84.33%) and the water content was a function of temperature; the water content increased with increase in temperature. Release studies of the ovalbumin from the agarose hydrogel nanoparticles revealed a diffusion-controlled release mechanism with a temperature dependence; the ovalbumin release rate was higher at 37 degrees C than that at room temperature. The great biocompatibility of agarose hydrogel, plus the mild conditions for drug encapsulation, make the agarose hydrogel nanoparticles a potential system for protein and peptide drug delivery.

Seroprevalence of hepatitis C virus infection and its genotype in Lanzhou, western China.

The seroprevalence of hepatitis C virus (HCV) infection in Lanzhou, Western China was studied. HCV genotypes in 20 patients with HCV infection was determined by genotype-specific primer for polymerase chain reaction (PCR) based on HCV core region and compared with the genotype assigned by sequence comparison and molecular evolutionary analysis based on the same region. Antibody to HCV (anti-HCV) was present in 2.5% of volunteer blood donors and in 35.0% of paid blood donors (P < 0.01). HCV infection is uncommon in patients with liver disease who attended liver clinics in this locality; 4.0% with acute hepatitis and 4.0% with chronic hepatitis, 10.0% with liver cirrhosis, and none with hepatocellular carcinoma were seropositive for anti-HCV. Genotype 1b and 2a were both found to be prevalent. Together, they accounted for 19 of 20 (95%) patients with HCV infection. Sequencing of the HCV core region from two patients showed that the assignment of HCV genotype by genotype-specific primers for PCR matched well with the genotyping results based on sequence comparison and molecular evolutionary analysis. These data showed that HCV is present in Western China, HCV infection is more common in paid blood donors, and HCV genotypes 1b and 2a are both prevalent in Western China.

Genetic analysis of the DNA recognition sequence of the P2 Cox protein.

The Cox protein of temperate Escherichia coli phage P2 is involved in three important biological processes: (i) excision of the integrated prophage genome (G. Lindahl and M. Sunshine, Virology 49:180-187, 1972), (ii) transcriptional repression of the P2 Pc promoter, which controls the expression of the immunity repressor C and the integrase (S. Saha, E. Haggård-Ljungquist, and K. Nordström, EMBO J. 6:3191-3199, 1987), and (iii) transcriptional activation of the late PII promoter of the unrelated satellite phage P4 (S. Saha, E. Haggård-Ljungquist, and K. Nordström, Proc. Natl. Acad. Sci. USA 86:3973-3977, 1989). A comparison of the DNA regions protected by Cox from DNaseI degradation has revealed a presumptive Cox recognition sequence (Saha et al., Proc. Natl. Acad. Sci. USA). The binding region of Cox in the P2 Pc promoter contains three presumptive recognition sequences, "Cox boxes," located in tandem. P2 vir3 and P2 vir24 are virulent deletion mutants unable to plate on Cox-producing strains, most likely because the deletions locate the new early promoters too close to the Cox-binding region (Saha et al., EMBO J.). In this report, spontaneous P2 vir3 and vir24 mutants, no longer sensitive to repression by the Cox protein, have been isolated. These mutants plate with equal efficiency on strains with or without a Cox-producing plasmid, and they have been named cor for cox resistance. Three types are recognized; the four P2 vir3 cor mutants have a 1-base deletion in the first Cox box, while the P2 vir24 cor mutants were of two types; four have a base substitution in the first Cox box, and one has a base substitution in the second Cox box. The effect of the Cox protein on the mutated P2 vir3 and vir24 promoters was analyzed in vivo by using fusions to a promoterless cat (chloramphenicol acetyltransferase) gene. The activities of the P2 vir3 and vir24 early promoters, as opposed to the wild-type early Pe promoter, are drastically reduced by the Cox protein, and the cor mutation renders them as resistant to Cox as the wild-type Pe promoter. Thus, at least the first two Cox boxes are essential for binding of the Cox protein.