[Analysis of risk factors associated with acute Stanford type B aortic dissection complicated with pleural effusion and observation of the curative effect after intracavitary repair].

Objective: To investigate the risk factors of acute Stanford type B aortic dissection (TBAD) complicated with pleural effusion (PE) and the short-term and long-term outcomes of thoracic endovascular aortic repair (TEVAR). Methods: A case-control study. The clinical and imaging data of 1 083 patients with acute TBAD admitted to the General Hospital of Northern Theater Command from April 2002 to December 2020 were retrospectively analyzed, including 211 cases with pleural effusion and 872 cases without pleural effusion. The baseline analysis of the two groups of patients was performed. The risk factors associated with pleural effusion were analyzed by binary logistic regression, and the results were expressed as odds ratio (OR) and 95% confidence interval (CI). According to the quantity of pleural effusion, they were simultaneously divided into small pleural effusion group and medium large pleural effusion group, to compare the short-term and long-term effects of TEVAR patients with different amounts of pleural effusion. Results: The incidence of pericardial effusion (17.5% vs. 3.8%, P<0.001), anemia (21.3% vs. 12.5%, P=0.001), aortic spiral tear (49.8% vs. 37.8%, P=0.002), dissection tear over diaphragm (57.8% vs. 48.1%, P=0.011), serum creatinine [85 (69, 111) vs. 81 (67, 100) µmol/L, P=0.011] and white blood cell levels[(11.3±4.2)×109/L vs. (10.3±4.2)×109/L, P=0.002] in acute TBAD pleural effusion group were significantly higher than those in non-pleural effusion group, and the hemoglobin level was significantly lower than that in non-pleural effusion group [(128±20) vs. (133±17) g/L, P<0.05]. Logistic stepwise regression analysis showed that pericardial effusion (OR=5.038,95%CI 2.962-8.568,P<0.001), anemia (OR=2.047,95%CI 1.361-3.079,P=0.001), spiral tear (OR=1.551,95%CI 1.030-2.336, P=0.002) and elevated white blood cell (OR=1.059,95%CI 1.011-1.102, P=0.005) were independent risk factors for TBAD complicated with pleural effusion. The incidences of all-cause death (4/19 vs. 1.5% vs. 0.9%, P<0.001), aortogenic death (4/19 vs. 0.7% vs. 0.7%, P<0.001) and aortic related adverse events (4/19 vs. 1.5% vs. 1.1%, P<0.001) in patients with large pleural effusion during TEVAR operation were significantly higher than those in patients with small pleural effusion and those without pleural effusion, and the differences were statistically significant. At 1 month follow-up after TEVAR, the incidence of all-cause death (4/16 vs. 3.3% vs. 1.6%, P<0.001), aortogenic death (4/16 vs. 0.8% vs.0.7%, P<0.001), aorta related adverse events (4/16 vs. 4.1% vs. 4.7%, P=0.013) and overall clinical adverse events (4/16 vs.9.8% vs. 6.7%, P=0.014) in the medium and large thoracic group were significantly higher than those in the small pleural effusion group and no pleural effusion group, and the differences were statistically significant. At 1 year follow-up after TEVAR, the incidence of all-cause death (4/15 vs. 4.9% vs. 3.9%, P=0.004), aortogenic death (4/15 vs.2.5% vs. 2.1%, P<0.001), aorta related adverse events (5/15 vs. 11.5% vs. 9.4%, P=0.012) and overall clinical adverse events (5/15 vs. 18.9% vs. 13.1%, P=0.029) in the medium and large thoracic group were significantly higher than those in the small pleural effusion group and no pleural effusion group, and the differences were statistically significant. Conclusions: Single center data showed that pericardial effusion, anemia, spiral tear and elevated white blood cell were independent risk factors for acute TBAD complicated with pleural effusion; the early (1 month) and long-term (1 year) rates of all-cause death, aortic mortality, aortic adverse events and overall clinical adverse events were significantly higher in TBAD patients with moderate pleural effusion after TEVAR, and moderate and large pleural effusion was an independent risk factor for near and long-term aortic related adverse events after TEVAR surgery.

[One-year follow-up results of atrial fibrillation patients who undergoing transcatheter aortic valve implantation].

Objective: To investigate whether atrial fibrillation (AF) before transcatheter aortic valve implantation (TAVI) will affect the prognosis of patients post TAVI. Methods: This is a single center retrospective study. A total of 115 patients with severe aortic stenosis (AS) who were admitted to General Hospital of Northern Theater Command from May 2016 to November 2020 and successfully received TAVI treatment were included. According to absence or accompaniment of AF pre-TAVI, they were divided into AF group (21 cases) and non-AF group (94 cases). The patients were followed up for postoperative antithrombotic treatment and the occurrence of the net adverse clinical and cerebrovascular events (NACCE) at 12 months post TAVI, including cardiogenic death, readmission to hospital for heart failure, nonfatal myocardial infarction, ischemic stroke and severe bleeding (BARC levels 3-5). Univariate logistic regression was used to analyze the related factors of NACCE. Results: Among the 115 selected patients, age was (73.8±6.9) years, there were 63 males. And 21 cases (18.2%) were diagnosed as AFbefore TAVI. In terms of postoperative antithrombotic therapy, 48.9% (46/94) of the patients in the non-AF group received monotherapy and 47.9% (45/94) received dual antiplatelet therapy. In the AF group, 47.6% (10/21) received anticoagulants and 33.3% (7/21) received dual antiplatelet therapy. The proportion of patients in the AF group taking non-vitamin K antagonist oral anticoagulants (NOAC) was higher than that in the non-AF group (38.1% (8/21) vs. 2.1% (2/94), P<0.001). Patients in both groups were followed up to 12 months after TAVI. During the 12 months follow-up, the incidence of NACCE after TAVI was 14.3% (3/21) in the AF group, which was numerically higher than that in the non-AF group (6.4% (6/94)), but the difference was not statistically significant (P=0.441). The incidence of severe bleeding was significantly higher in the AF group than in the non-AF group (9.5% (2/21) vs. 0, P=0.032). Univariate logistic regression analysis showed that hypertension was associated with the risk of NACCE (OR=8.308, P=0.050), while AF was not associated with the risk of NACCE (P=0.235). Conclusion: The incidence of severe bleeding after TAVI is higher in patients with AF than in patients without AF prior TAVI, and there is a trend of increased risk of NACCE post TAVI in AF patients.

Clinician perspectives on a culturally adapted patient decision aid concerning maintenance therapy for asthma.

BACKGROUND AND OBJECTIVE: Patients with persistent asthma often show poor adherence to inhaled corticosteroids (ICS). Shared decision-making can improve adherence rates in this population. Patient decision aids (PDAs) are tools to facilitate shared decision-making. To date, only one PDA, developed in a Canadian French-speaking population, exists for patients considering ICS maintenance therapy. This PDA has been culturally adapted in this study to contextualize to the needs of multi-ethnic Asian patients in Singapore. This study explored the views of local clinicians on the content, design and implementation of this newly-adapted PDA. METHODS: 24 clinicians, who were purposively sampled from polyclinics and a tertiary institution, were interviewed on the content, design and implementation of the PDA. The interviews were audio-recorded, transcribed and analyzed via thematic analysis. RESULTS: Clinicians generally accepted the design of the PDA. They suggested for the target users to be patients on Step 2 of GINA guidelines and the number of options to be reduced from four to two (do nothing or start inhaled corticosteroids). Moreover, they supported including a list of values for patients to select from given that Asian patients often do not articulate their values readily. The addition of more visual aids, the production of multilingual Asian editions and the involvement of nurses to administer the PDA was also suggested. CONCLUSION: The PDA was culturally-adapted with local clinicians' perspectives to target multi-ethnic Asian patients with persistent asthma (Step 2 GINA guidelines). The main changes include a list of values and addition of visual aids.

[Perioperative serious complications in patients undergoing transcatheter aortic valve replacement: a single-center experience].

Objective: To observe the incidence of perioperative severe complications in aortic stenosis (AS) patients undergoing transcatheter aortic valve implantation (TAVR), and to explore the influence of the accumulated experience of the operators on the incidence of complications. Methods: This is a single-center retrospective study. Patients with AS who underwent TAVR from May 2016 to November 2020 in General Hospital of Northern Theater Command were included. The occurrence of severe complications during perioperative period was recorded. Severe complications included all-cause death, surgical transfer to thoracotomy, coronary artery occlusion, peripheral vascular approach complications, severe cerebrovascular events, moderate or above perivalvular leakage, valve displacement (implanted valve middle valve), pacemaker implantation, etc. In order to observe the influence of the accumulated experience of the operators on the occurrence of postoperative complications, the complications in each stage of the patients were counted and the bar chart was drawn with interval of every 30 patients. Results: A total of 119 patients were included, including 64 males and 55 females, the mean age was (73.9±6.9) years. The valve implantation was unsuccessful in 3 out of 119 patients (2.5%). There were 39 cases of severe complications during perioperative period, including 1 death (0.8%), 2 cases of thoracotomy (1.7%), 2 cases of coronary artery occlusion (1.7%), 8 cases of peripheral vascular approach complications (6.7%), and 1 case of new severe cerebrovascular event (0.8%), 3 cases of moderate or higher perivalvular leakage (2.5%), 8 cases of valve displacement leads to midvalvular implantation (6.7%), 14 cases of permanent pacemaker implantation due to new atrioventricular block (11.8%). The bar chart showed that the incidence of permanent pacemaker implantation and in-valve implantation did not significantly decrease with the accumulation of experience, while the incidence of other complications showed a decreasing trend after 30 cases. Conclusions: Most serious complications occurred in the early stage of TAVR in our center. The incidence of all cause death, coronary artery occlusion and peripheral vascular approach complications in the perioperative period post TAVR could be reduced in the TAVR center in the learning stage through the accumulation of procedure-related experience, but the incidence of pacemaker placement and the implanted valve within the valve dose not significantly decrease over time.

[The long coding RNA GSTM3TV2 acts an oncogene to promote chemoresistance in pancreatic cancer].

Objectives: To examine the expression of the long coding RNA GSTM3TV2 in pancreatic cancer tissues and to examine its role and mechanism in chemoresistance of pancreatic cancer cells. Methods: The expression of lncRNA GSTM3TV2 in 15 pancreatic cancer specimens and corresponding adjacent to cancer tissue samples diagnosed by Department of Pathology, Peking Union Medical College Hospital was detected by real-time PCR.And the expressions of GSTM3TV2 in pancreatic cancer cell AsPC-1, BxPC-3, MIAPaCa-2, PanC-1, SU86.86, T3M4, and chemoresistant cells AsPC-1/GR and MIAPaCa-2/GR, and human pancreatic nestin-expressing cells hTERT-HPNE were detected. Pancreatic cancer cell lines were transfected with GSTM3TV2-pcDNA3.1(+)in order to get cells with GSTM3TV2 overexpression.GSTM3TV2-siRNA was transfected into pancreatic cancer cells to knock down GSTM3TV2. The cell chemoresistance was measured by CCK-8 and flow cytometry assay when incubated with nab-paclitaxel. At the same time, subcutaneous xenograft tumor models were established in nude mice to observe the effect of GSTM3TV2 on chemoresistance of tumor growth in nude mice.Western blot assay was also performed to detect the molecular mechanism of chemoresistance of GSTM3TV2. Results: Comparing toadjacent tissues(0.084±0.019), GSTM3TV2 expression was significantly upregulated in the pancreatic cancer tissues(0.493±0.084) (t=5.146, P<0.05). GSTM3TV2 expression were higher in the chemotherapy resistance pancreatic cancer cells AsPC-1/GR(210.799±19.788) and MIAPaCa-2/GR(122.408±23.419) than that in the AsPC-1(3.793±0.615) and the MIAPaCa-2(5.179±1.095)(t=21.800,P<0.05;t=-18.490,P<0.05). The results of in vivo experiments showed that the volume of subcutaneously transplanted tumors in the overexpressing GSTM3TV2 group ((1 059.609±102.498)mm(3)) was significantly larger than that in the control group((566.414±81.087) mm(3)) by treated with nab-paclitaxel(t=4.230,P<0.05).Meanwhile, GSTM3TV2 could promote the expression of Cyclin D1, CDK6, Cyclin E1, Vimentin, N-cadherin, ZEB1, Snail and Slug; but decrease cleaved caspase-3, cleaved PARP in pancreatic cancer cells. Conclusions: The expression level of GSTM3TV2 in pancreatic canceris higher than that in paired adjacent tissues. GSTM3TV2 may act as an oncogene to promote chemoresistance in pancreatic cancer through regulation of cell proliferation, apoptosis, and epithelial-mesenchymal transition.

Asthma care based on Chronic Care Model in an aging Asian community.

To cope with the higher prevalence of asthma and other non-communicable diseases without compromising on quality of care, a Singapore public primary care institution has adopted the Chronic Care Model (CCM). This retrospective cohort study aimed to describe the proportion of patients with well-controlled asthma (based on Asthma Control Test score ≥20) between 2010 and 2016 in association with their management based on the CCM (which covers the polyclinic clinical information system, self-management measures, system re-design and decision support). Data were retrieved from the Singapore National Asthma Programme (SNAP) and institutional clinical quality databases of eight local polyclinics in eastern and southern Singapore. The data were aggregated, analysed and presented in proportions against monthly polyclinic attendances for asthma. From 2010 to 2016, the total asthma attendances increased by 31% from 27,345 to 35,731, with the highest rise among patients aged ≥60 years. The proportion of patients with good asthma control rose from 71.4% to 80.9%; those who received rescue therapy for acute exacerbations fell from 15.8% to 11.7% and those referred to emergency departments after failed rescue therapy decreased from 0.7% to 0.6%. The proportion of patients with updated asthma action plans increased from 66.7% to 73.4% (proxy for self-management). The overall health and process outcomes of asthma seemed to have improved with multiplex of system-based interventions relating to the introduction of CCM in a public primary healthcare institution in Singapore.

Rasagiline, an inhibitor of MAO-B, decreases colonic motility through elevating colonic dopamine content.

BACKGROUND: Dopamine (DA) is a negative modulator of gut motility. Monoamine oxidase-B (MAO-B) is an important metabolic enzyme degrading DA. Rasagiline, an irreversible MAO-B inhibitor, is used to treat Parkinson's disease because of its neuroprotective effect and increasing central DA. However, it is unclear whether MAO-B exists in the colon and rasagiline increases colonic DA, thereby affecting colonic motility. METHODS: Immunohistochemistry, western blotting, enzyme activity assay, colonic motility recording, gut transit test, and high-performance liquid chromatography-electrochemical detection were employed in this study. KEY RESULTS: Monoamine oxidase-B was distributed in the colonic muscular layers including neurons and glias of rat and human. When oral treatment of rats with rasagiline for 4 weeks, in vitro colonic motility was significantly reduced, but it was greatly reversed by SCH-23390, an antagonist of DA D1 receptor. The rasagiline-treated rats also manifested decreased MAO-B activity and increased DA content in the colonic muscular layer, but no alterations were detected in the protein expressions of D1 and D2 receptors, and MAO-A and MAO-B, as well as in the content of 5-hydroxytryptamine and noradrenaline. Moreover, acute administration of rasagiline did not affect the colonic motility in vitro and the colonic DA level in rats, although MAO-B activity was significantly inhibited. CONCLUSIONS & INFERENCES: Monoamine oxidase-B is abundant in the colonic muscular layer including myenteric plexus of rat and human. Long-term administration of rasagiline can increase colonic DA thereby inhibiting colonic motility, suggesting that colonic MAO-B could be a potential drug target for colonic dysmotility.

[The expression of KLK7 in pancreatic cancer and the effects on the biological behavior of pancreatic cancer cells].

Objective: To investigate the expression of KLK7 in pancreatic cancer and its clinical significance. Methods: Immunohistochemistry was used to detect the expression of KLK7 protein in pancreatic cancer tissue microarray with 92 samples. Statistical analysis of the relationship between KLK7 and clinicopathological characteristics was finished. Pancreatic cancer cell lines were infected with lentiviuses in order to get cells with KLK7 stable overexpression.KLK7-siRNA was transfected into pancreatic cancer cells to knock down KLK7.Cell proliferation and chemosensitivity were detected by CCK-8 assay; Cell invasion and migration abilities were detected by Transwell assay. At the same time, subcutaneous xenograft tumor models were established in nude mice to observe the effect of KLK7 on tumor growth in nude mice. Data were statistically analyzed by rank sum test, χ(2) test and Logistic regression analysis. Results: The expression level of KLK7 in pancreatic cancer tissues was higher than that in paired adjacent tissues (P<0.05). KLK7 expression was correlated with vascular invasion(χ(2)=7.535, P<0.05). Further univariate and multivariate analysis showed that KLK7 expression was an independent risk factor for vascular invasion of pancreatic cancer(χ(2)=7.535, P<0.05). The overexpression of KLK7 in pancreatic cancer cell lines BxPC-3 and CFPAC can increase their proliferation abilities, reduce the chemosensitivity and promote their migration and invasion behaviour; The results of in vivo experiments showed that the volume of subcutaneously transplanted tumors in the overexpressing KLK7 group was significantly larger than that in the control group (t=4.479, P<0.05). The group of overexpressing KLK7 showed greater tumor weight than the control group(t=2.831, P<0.05). Conclusions: The expression level of KLK7 in pancreatic ductal adenocarcinoma was higher than that in paired adjacent tissues and it is an independent risk factor for vascular invasion of pancreatic cancer.KLK7 can promote the proliferation of pancreatic cancer cells, reduce the chemosensitivity and increase the invasion and migration of pancreatic cancer cells.

[A prospective control study of Saccharomyces boulardii in prevention of antibiotic-associated diarrhea in the older inpatients].

Objective: To study the value of Saccharomyces boulardii for the prevention of antibiotic-associated diarrhea in older inpatients. Methods: A total of 163 older patients who were treated with wide-spectrum antibiotics at least three days during January 2014 to December 2015 were randomly divided into control and study group. In study group, 81 patients were administrated with oral Saccharomyces boulardii 500 mg twice a day for 21 days. The control group was of no intervention. Morbidity rate of antibiotic-associated diarrhea and Clostridium difficile-associated diarrhea, frequency and duration of diarrhea were recorded. Results: The incidence of antibiotic-associated diarrhea in study group was significantly lower than that in control group [14.8%(12/81) vs 28.0%(23/82), P<0.05], whereas no difference was seen in the incidence of Clostridium difficile-associated diarrhea [3.7%(3/81) vs 4.9%(4/82), P>0.05] in two groups. The frequency and duration of diarrhea in the study group were significantly lower and shorter than those in control group[(4.3±1.7) times/day vs (6.9±2.0) times/day; (3.0±1.1) days vs (5.7±1.8) days, both P<0.01]. Conclusion:Saccharomyces boulardii may reduce the incidence of antibiotic-associated diarrhea therefore improving the symptom of diarrhea in older inpatients.

[Correlation between miR-1178 expression and clinicopathological significance in human pancreatic cancer].

Objective: To test the expression of miR-1178 in pancreatic cancer and study its clinicopathological significance and mechanism. Methods: The expression of miR-1178 in 87 paired paraffin pancreatic ductal adenocarcinoma specimens and adjacent non- cancerous pancreatic tissue diagnosed by Pathology Department of Peking Union Medical College Hospital was detected by hybridization in situ. The relationship between the expression of miR-1178 and clinicopathological characters was analyzed.miR-1178 mimics and inhibitor were used to further detect the close relationship among miR-1178 and cancer invasion. Establish a nude mice subcutaneously transplanted tumor model, 4 weeks after vaccination for tumor volume and weight measurement.Student t-test, rank sum test, and χ(2) test was used respectively to compare the data between two groups. Cox regression was adopted to improve the single factor and multiple factors analysis. Results: The results of hybridization in situ showed the expression of miR-1178 was increased in 72 cases with pancreatic cancer compared to that in paired normal pancreatic tissues (50/72 vs. 11/72, χ(2)=43.26, P<0.05). miR-1178 expression was positively associated with tumor lymph node stage (χ(2)=4.189, P=0.041). Univariate and multivariate analysis revealed that miR-1178 was an independent adverse prognostic indicator for patients with pancreatic cancer (HR=2.364, 95%CI: 1.114-5.019, P=0.025). Transwell assay indicated the over-expression of miR-1178 increased the number of AsPC-1 cells that penetrated the ECM-coated membrane (177.0±19.8 vs. 119.7±15.9)(χ(2)=8.21, P<0.05). For the in vivo experiment, overexpression of miR-1178 significantly promoted tumor growth, compared with control group (tumor volume: (5 122.4±760.2)mm(3) vs. (1 976.8±601.8)mm(3), t=2.413, P<0.05; tumor weight: (1.55±0.21)g vs. (0.67±0.17)g, t=2.960, P<0.05). Over-expression of miR-1178 down-regulated the expression of Stub1 and elevated the expression of FAK/MMP-9 signal pathway(P<0.05). Conclusions: MiR-1178 is overexpressed in pancreatic cancer, and is effective for predicting patients' prognosis. MiR-1178 regulate Stub1/FAK/MMP-9 signal pathway and promote the invasion of AsPC-1 cells.

The role of the vagal pathway and gastric dopamine in the gastroparesis of rats after a 6-hydroxydopamine microinjection in the substantia nigra.

AIM: Gastroparesis is a common non-motor system symptom of Parkinson's disease (PD). However, the mechanism responsible for the gastric motor abnormality is not clear. We previously reported on the impaired gastric motility in 6-hydroxydopamine (6-OHDA) rats, which were treated with a bilateral microinjection of 6-OHDA in the substantia nigra (SN). We hypothesize that the enhanced dopamine system and reduced acetylcholine (Ach) in gastric tissues might contribute to the delayed gastric emptying observed in PD. METHODS: A strain gauge force transducer, digital X-ray imaging system, Western blot, immunofluorescence and Radio Immunoassay were used in this study. RESULTS: Dopaminergic neurones in the SN were greatly reduced following the bilateral microinjection of 6-OHDA. 6-OHDA rats exhibited impaired gastric motility and delayed gastric emptying, accompanied by increased dopamine content and the overexpression of D2 receptors in the stomach. The administration of the D2 receptor antagonist domperidone relieved gastric dysmotility in 6-OHDA rats, but the D1 receptor antagonist SCH23390 failed to do so. Subdiaphragmatic vagotomy prevented the increase in the gastric dopamine content and D2 receptor expression and improved gastric dysmotility in 6-OHDA rats. CONCLUSION: Dopaminergic deficiency in the SN results in impaired gastric motility, possibly as a result of the enhanced activity of dopamine system and reduced Ach in gastric tissue. The vagus nerve plays an important role in peripheral gastric motility disorder.

Effects of konjac flour inclusion in gestation diets on the nutrient digestibility, lactation feed intake and reproductive performance of sows.

This study was conducted to investigate the effects of konjac flour (KF) inclusion in gestation diets of sows on nutrients digestibility, lactation feed intake, reproductive performance of sows and preweaning performance of piglets. Two isoenergetic and isonitrogenous gestation diets were formulated: a control diet and a 2.1% KF-supplemented diet (KF diet). Both diets had the same NDF and insoluble fiber (ISF) levels, but the KF diet had higher soluble fiber (SF) level. The day after breeding, 96 multiparous sows were assigned to the two dietary treatments. Restrict-fed during gestation, in contrast, all sows were offered the same lactation diet ad libitum. Response criteria included sow BW, backfat depth, lactation feed intake, weaning-to-estrus interval, litter size and piglet's weight at parturition and day 21 of lactation. On day 60 of gestation, 20 sows were used to measure nutrient digestibility. Results showed that the digestibility of dry matter, gross energy, crude fiber and ADF were not affected by the dietary treatments. The inclusion of KF in gestation diets increased NDF digestibility (P<0.05) and tended to increase the digestibility of CP (P=0.05) compared with the control diet group. In addition, dietary treatment during gestation did not affect litter size, BW and backfat gain during gestation, lactation weight, backfat loss or weaning-to-estrus interval of sows. However, sows fed the KF diet consumed more (P<0.05) lactation diet per day than sows in the control group. Accordingly, sows fed the KF diet showed greater average piglet weights on day 21 of lactation (P=0.09), and the litter weight of sows fed the KF diet on day 21 of lactation increased by 3.95 kg compared with sows fed the control diet (not significant). In conclusion, the inclusion of KF in gestation diets increased lactation feed intake of sows and tended to improve litter performance.

Cellular distribution of NKCC2 in the gastric mucosa and its response to short-term osmotic shock.

The Na(+)-K(+)-2Cl(-) cotransporter-2 (NKCC2) has long been recognized as a "kidney-specific" transporter and is important in salt reabsorption. NKCC2 has been found in the gastric mucosa; however, its cellular distribution and function remain obscure. The present study characterized the distribution pattern of NKCC2 in mammalian gastric mucosa and investigated its response to osmotic challenge. Reverse transcription with the polymerase chain reaction, Western blot and immunofluorescence were used to determine NKCC2 expression and localization. The effect of osmotic shock on NKCC2 expression was studied in isolated gastric mucosa with variable osmolarity treatment. Results from all of the above studies were compared with those of NKCC1. Our data indicated that NKCC1 and NKCC2 were expressed in the gastric mucosa of rat, mouse and human. The mRNA transcripts and proteins for NKCC1 and NKCC2 were broadly expressed in the rat gastric mucosa. In rat and mouse, NKCC1 was largely confined to the lower part of the oxyntic and pyloric gland areas, whereas NKCC2 extended throughout the gastric glands. NKCC1 immunoreactivity was strongly expressed in the parietal and chief cells but was weaker in the mucous cells. NKCC2 was abundantly located in the parietal and mucous cells but faintly distributed in the chief cells. Hypertonic treatment increased the protein level of NKCC1 and caused evident membrane translocation. In contrast, NKCC2 was significantly downregulated and no obvious membrane translocation was observed. Thus, NKCC2 displayed a more ubiquitous distribution in the gastric mucosa and might work coordinately with NKCC1 to maintain cell volume homeostasis under hypertonic conditions.

Entacapone promotes cAMP-dependent colonic Cl(-) secretion in rats.

BACKGROUND: Entacapone is a promising drug used widely for the treatment of Parkinson's disease (PD) as a catechol-O-methyl transferase (COMT) inhibitor. However, entacapone has gastrointestinal side effects. The aim of this study was to investigate the effects of entacapone on the epithelial ion transport in rat distal colon, and explore the underlying mechanism. METHODS: The study was performed on freshly isolated colonic mucosa-only, submucosa-only and mucosa-submucosa preparations in rat. The short circuit current (I(SC) ) was measured to determine electrogenic ion transport, and a scanning ion-selective electrode technique (SIET) was used to directly measure Cl(-) flux across the epithelium. The content of intracellular cAMP was measured with radioimmunoassay (RIA). KEY RESULTS: Entacapone increased mucosal I(SC) in the rat distal colon. I(SC) was inhibited significantly by apical addition of diphenylamine-2,2'-dicarboxylic acid (DPC), a blocker of the Cl(-) channel, basolateral application of bumetanide, an inhibitor of Na(+) -K(+) -2Cl(-) co-transporter (NKCC), removal of Cl(-) from the bathing solution, and pretreatment with MDL 12330A, an inhibitor of adenylate cyclase. Inhibiting endogenous prostaglandin (PG) synthesis with indomethacin, and eliminating submucosal enteric neural activity with tetrodotoxin (TTX)-inhibited entacapone-evoked I(SC) increases. Similar results were also obtained when Cl(-) flux was measured with SIET. Entacapone significantly increased intracellular cAMP content, which was greatly inhibited by either indomethacin or TTX in the tissues containing submucosal plexus, and by only indomethacin in the mucosa-only preparations. CONCLUSIONS & INFERENCES: Entacapone stimulates cAMP-dependent Cl(-) secretion in the rat colon, and this process is regulated by endogenous PG and the submucosal enteric nervous system.

Expression and activation of ß-adrenoceptors in the colorectal mucosa of rat and human.

BACKGROUND: The functions of the distal colon are regulated by local and extrinsic neural pathways. In previous studies, we have found that dopamine (DA) and norepinephrine (NE) could evoke colonic ion transport by activating ß-adrenoceptors. The present study aims to investigate the segmental differences in expression and activation of ß-adrenoceptors in the distal colon in physiological and pathophysiological conditions. METHODS: Real-time PCR, immunofluorescence, and Western blotting were used to detect the expression of ß-adrenoceptors in the rat and human distal colon. Short-circuit current measurements (Isc) were used to assess the role of ß-adrenoceptors in ion transport. KEY RESULTS: DA and NE caused greater suppression of baseline Isc in distal colon adjacent to the rectum than in segments further away from the anus. These responses were inhibited by selective antagonists of ß1- and ß2-adrenoceptors, but not ß3-adrenoceptor. The expression levels of ß1- and ß2-adrenoceptors in colonic mucosa were higher in colorectum than the regions away from the anus of rats and humans. In wrap-restraint stress (2 h), DA-, NE-induced ΔIsc and the expression of ß-adrenoceptors in the colorectum were significantly reduced. However, when endogenous catecholamines were depleted by 6-hydroxydopamine (75 mg kg(-1), i.p., 3 days), DA-, NE-induced ΔIsc as well as the expression of ß-adrenoceptors were significantly enhanced in the rat colorectum but not in more proximal regions of the distal colon. CONCLUSIONS & INFERENCES: ß1- and ß2-adrenoceptors are predominantly expressed in the colorectal mucosa. Perturbation of endogenous catecholamine levels influences the expression and activation of ß-adrenoceptors in the colorectal region.

Activation of submucosal 5-HT(3) receptors elicits a somatostatin-dependent inhibition of ion secretion in rat colon.

BACKGROUND AND PURPOSE: 5-Hydroxytryptamine (5-HT) is a key regulator of the gastrointestinal system and we have shown that submucosal neuronal 5-HT(3) receptors exerted a novel inhibitory effect on colonic ion transport. The aim of the present study was to investigate the precise mechanism(s) underlying this inhibitory effect. EXPERIMENTAL APPROACH: Mucosa/submucosa or mucosa-only preparations from rat distal colon were mounted in Ussing chambers for measurement of short-circuit current (I(sc)) as an indicator of ion secretion. Somatostatin release was determined with radioimmunoassay. Intracellular cAMP content was measured with enzyme-linked immunoadsorbent assay (elisa). Immunohistochemical techniques were used to study the expression of 5-HT(3) receptors, somatostatin and somatostatin receptors in colonic tissue. KEY RESULTS: In rat distal colonic mucosa/submucosa preparations, pretreatment with 5-HT(3) receptor antagonists enhanced 5-HT-induced increases in I(sc). However, in mucosa-only preparations without retained neural elements, pretreatment with 5-HT(3) receptor antagonists inhibited 5-HT-induced DeltaI(sc). Pretreatment with a somatostatin-2 (sst(2)) receptor antagonist in mucosa/submucosa preparations augmented 5-HT-induced DeltaI(sc). Combination of sst(2) and 5-HT(3) receptor antagonists did not cause further enhancement of 5-HT-induced DeltaI(sc). Moreover, both sst(2) and 5-HT(3) receptor antagonists enhanced 5-HT-induced increase in intracellular cAMP concentration in the mucosa/submucosa preparations. 5-HT released somatostatin from rat colonic mucosa/submucosa preparations, an effect prevented by pretreatment with 5-HT(3) receptor antagonists. Immunohistochemical staining demonstrated the presence of 5-HT(3) receptors on submucosal somatostatin neurons and of sst(2) receptors on colonic mucosa. CONCLUSION AND IMPLICATIONS: Activation of neuronal 5-HT(3) receptors in the submucosal plexus of rat colon suppressed 5-HT-induced ion secretion by releasing somatostatin from submucosal neurons.

Expression of NKCC2 in the rat gastrointestinal tract.

NKCC2, an isoform of Na+-K+-2Cl(-) cotransporter, is principally present in the kidney and plays a critical role in salt reabsorption. Expression of NKCC2 has been found in the apical membrane of intestinal epithelial cells in a number of marine fish, however, details for expression in the mammalian gastrointestinal tract are lacking. RT-PCR, Western blotting and immunohistochemistry were used to study the expression and localization of NKCC2 in the rat gastrointestinal tract. We found that mRNA transcripts, protein and immunoreactivity (IR) for NKCC2 were expressed in the stomach, small and large intestine of adult rats. NKCC2 IR was localized to the base of the gastric glands, intestinal epithelia, myenteric and submucosal plexuses. NKCC2 IR was expressed strongly in the apical membranes and weakly in the basolateral membranes of intestinal epithelial cells. In the enteric nervous system, NKCC2 IR was widely distributed and localized to enteric neurons with cholinergic, calretinin and nitrergic neuronal immunochemical codes in the myenteric plexus. It was localized to non-cholinergic secretomotor neurons in the submucosal plexus. In conclusion, this study for the first time clearly detected the expression of NKCC2 in the gastrointestinal tract of a mammalian species. Expression of NKCC2 in gastrointestinal epithelial cells suggested that this cation chloride cotransporter might be involved in gastrointestinal ion transport. Expression of NKCC2 in enteric neurons might contribute to the accumulation of Cl(-) and a more depolarized E(Cl)(-) in enteric neurons.

Alteration of dopaminergic markers in gastrointestinal tract of different rodent models of Parkinson's disease.

Parkinson's disease (PD) is a progressive neurological disorder that is often associated with various gastrointestinal (GI) symptoms. The link between the alteration of dopaminergic system and the symptoms of the GI tract in PD is complicated. To determine the changes in the dopaminergic system in the GI tract in PD, two kinds of rodent PD models were used in the present study. One was 6-hydroxydopamine (6-OHDA) -treated rats in which 6-OHDA was microinjected in the bilateral substantia nigra (SN). The other was 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) -treated mice in which MPTP was injected intraperitoneally. Immunofluorescence, reverse transcription (RT)-real time polymerase chain reaction (PCR) and Western blot were used to evaluate and compare the levels of mRNA and protein expression of tyrosine hydroxylase (TH) and dopamine transporter (DAT) in the GI tract between normal and rodent PD models, as well as between 6-OHDA-treated rats and MPTP-treated mice. The results indicated that TH- and DAT-positive cells were widely distributed in the GI tract. There were significant differences in TH and DAT expression in the GI tract between normal and PD models, as well as between 6-OHDA-treated rats and MPTP-treated mice. The protein levels of TH and DAT in the GI tract were significantly increased in 6-OHDA-treated rats, but the protein level of TH was significantly decreased in MPTP-treated mice. In addition, there was visible atrophy of gastric epithelial parietal cells in MPTP-treated mice, although the protein level of DAT was not significantly changed. The different alterations of dopaminergic system in the GI tract of the two kinds of PD models might underline the differences in GI symptoms in PD patients and might be correlated with the disease severity and disease process affecting the GI tract.

Pharmacokinetics of perlolyrine in rats by stable isotope dilution in conjunction with GC-MS.

AIM: To determine the pharmacokinetics of perlolyrine in rats. METHODS: The plasma concentration and pharmacokinetic parameters of perlolyrine were determined by gas chromatography-mass spectrometry (GC-MS) with selected ion (m/z 247 and m/z 248) and [2-(15) N] perlolyrine (m/z 248) as internal standard. RESULTS: The concentration-time profile of perlolyrine after ig perlolyrine 2 mg.kg-1 fitted a two-compartment open model in rats. The pharmacokinetic parameters were T1/2 alpha = 0.33 h, T1/2 beta = 4.52 h, T1/2 (ka) = 0.14 h, Tmax = 0.35 h, Cmax = 18.84 micrograms/L, K12 = 0.88 h-1, K21 = 0.42 h-1, K10 = 0.32 h-1, V/F = 109.22 L.kg-1, AUC = 112.68 micrograms.h.L-1. CONCLUSION: The method was constant, sensitive, and accurate. It provides a useful method for the determination of pharmacokinetics of perlolyrine which are important for clinical use of perlolyrine.

[Study on the organization of chromatin and chromosome in mouse spermatogenic cells by scanning electron microscopy].

It remains unclear about the intermediate construction of chromosome due to its highly compact nature and the limitation in methods. The present study was designed to investigate the construction of chromatin and mitotic chromosome in situ with scanning electron microscopy. Mouse testes were selected as the material, because of in which the spermatogenic cells divide actively and successively to form the sperm. Such a feature would be able to study the structure of mammalian chromatin and chromosomes along with the change of nuclear cycle. The animal were perfused with 200 ml of 0.075 mol/L KCl hypotonic solution to remove blood and placed for 15-20 min on ice followed by 0.5% glutaraldehyde and 0.5% formaldehyde for fixing. Through treated by the routine process of fractured and freeze dried with t-butyl alcohol, the specimens were then coated with a 3 nm thick platinum and observed with Hitachi S-430 scanning electron microscopy. It was found that the hypotonic treatment with 0.075 mol/L KCl solution was suit for demonstrating the nuclear structure, when the organelles were well preserved. The chromatin fibers of 10-30 nm and 80-125 nm in diameter could be recognized in the interphase nuclei, which were arranged losely at the region of euchromatin, and folded with each other into chromatin masses at the region of heterochromatin, while the chromatin fibers with the diameter of 80-125 nm often could be viewed on the mitotic chromosomes. Since its presence in interphase nuclei and mitotic chromosomes, it was considered that the chromatin fibers with 80-125 nm in diameter might play a role in the condensation of chromosome, serve as a type of the intermediate structure.