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Background: The gut is capable of inducing multiple organ dysfunction syndrome (MODS). In the diagnosis and treatment of critical ill patients, doctors should pay particular attention to the protection or recovery of intestinal barrier function. However, no reliable diagnostic criteria are available clinically. This study aimed to assess the changes of intestinal mucosal barrier function in surgically critical ill patients as well as their significance., Methods: Thirty-eight surgically critical ill patients were enrolled as a study group (APACHE II>8 scores), and 15 non-critical ill patients without intestinal dysfunction were selected as a control group (APACHE II<6). General information, symptoms, physical signs, and APACHE II scores of the patients were recorded. The patients in the study group were subdivided into an intestinal dysfunction group (n=26) and a non-intestinal dysfunction group (n=12). Three ml venous blood was collected from the control group on admission and the same volume of plasma was collected from the study group both on admission and in the period of recovery. The plasma concentrations of endotoxin, diamine oxidase (DAO), D-lactate, and intestinal fatty-acid binding protein (iFABP) were detected respectively. The data collected were analyzed by the SPSS 17.0 software for Windows., Results: The levels of variables were significantly higher in the study group than in the control group (P<0.01). They were higher in the intestinal dysfunction group than in the non-intestinal dysfunction group (DAO P<0.05, endotoxin, D-lactate, iFABP P<0.01). In the non-intestinal dysfunction group compared with the control group, the level of endotoxin was not significant (P>0.05), but the levels of DAO, D-lactate and iFABP were statistically significant (P<0.05). The levels of variables in acute stage were higher than those in recovery stage (P<0.01). The death group showed higher levels of variables than the survival group (endotoxin and D-lactate P<0.01, DAO and iFABP P<0.05)., Conclusion: The plasma concentrations of endotoxin, DAO, D-lactate, and intestinal fatty-acid binding protein (iFABP) could reflect a better function of the intestinal mucosa barrier in surgically critical ill patients.

Objective: To investigate the mechanism of ursodeoxycholic acid (UDCA) in hepatocyte apoptosis using differentiated hepatocytes derived from bone marrow mesenchymal cells., Methods: Rat bone marrow mesenchymal cell was induced into mature hepatocytes in vitro and then treated with PBS (Cont), deoxycholic acid (DCA), DCA plus UDCA (U + D) or UDCA alone (UDCA). Cell apoptosis was detected by Hoechst staining and Caspase-3 activity measurement. The mRNA expressions of p53 and Bax were measured by semi-quantitative RT-PCR and real-time RT-PCR. The Bax protein expression was detected by immunohistochemistry., Results: In comparison with Cont, DCA obviously induced hepatocyte apoptosis as measured by an increased cell count and a higher Caspase-3 activity (P < 0.05). These increments could be inhibited by addition of UDCA. The expressions of p53 and Bax in hepatocytes were up-regulated by DCA. These up-expressions could also be inhibited by UDCA. The DCA-induced increased count of Bax-positive cells could be reduced by UDCA., Conclusion: UDCA inhibits DCA-induced hepatocyte apoptosis by down-regulating the expression of p53/Bax signal molecule.

We have recently shown that membrane association of neuronal nitric oxide synthase-alpha (nNOSalpha) is critical in the regulation of synthesis of NO during nitrergic neurotransmission. The purpose of this study was to examine the role of the synapse-associated proteins (SAPs) in membrane association of nNOSalpha. Varicosities (swellings on terminal axons) were isolated from mice gastrointestinal tract and examined for nNOSalpha, postsynaptic density protein 95 (PSD95), and membrane interactions by coimmunoprecipitation and SDS-PAGE. Our results show that PSD95 protein was present in the membrane fraction of the nerve varicosity, whereas both PSD95 and SAP97 were present in the cytosol. nNOSalpha was associated with PSD95 but not SAP97. nNOSalpha-PSD95 complex was bound to the membrane via palmitoylation of PSD95. Depalmitoylation of PSD95 with 2-bromopalmitate dislocates nNOSalpha and PSD95 from the varicosity membrane and abolishes NO production. These studies show that palmitoylation of PSD95 anchors nNOSalpha to the varicosity membrane and that it is obligatory for NO production by the enzyme. Palmitoylation of PSD95 may provide a novel target for regulation of nitrergic neurotransmission.

Objective: To investigate the risk factors of intrahepatic cholangiocarcinoma (ICC) and extrahepatic cholangiocarcinoma (ECC)., Methods: The clinicopathological data of 190 patients with cholangiocarcinomas (61 ICC and 129 ECC) diagnosed and treated in the Peking Union Medical College Hospital between 1998 and 2008 were collected. The clinicopathological data of 380 matched healthy controls were also collected. The information about liver diseases, family history, diabetes, smoking and drinking were recorded and analyzed., Results: The positive rate of HBsAg(+) and anti-HBc(+), HBsAg(-) and anti-HBc(+) and the incidence of choledocholithiasis or hepatolithiasis in ICC patients were 27.9%, 50.8% and 14.8%, respectively. The incidence of diabetes mellitus, cholecystolithiasis, choledocholithiasis or hepatolithiasis and previous cholecystectomy in ECC patients were 18.6%, 15.5%, 18.6% and 13.2%, respectively. The incidences of all above mentioned factors in the ICC or ECC patients were significantly higher than that in the controls (P < 0.05). Compared with the patients with ECC, the ICC patients had a significantly higher cirrhosis rate (P < 0.05)., Conclusion: Our study results show that choledocholithiasis or hepatolithiasis, liver cirrhosis and chronic HBV infection are possible risk factors for intrahepatic cholangiocarcinoma, while choledocholithiasis or hepatolithiasis, diabetes mellitus, cholecystolithiasis, history of cholecystectomy are risk factors for extrahepatic cholangiocarcinoma.

Objective: To evaluate the surgical technique of "Kou mode of hepatic hilar anastomosis" in the treatment for type III or IV hilar cholangiocarcinoma., Methods: The clinical data of 89 patients with type III or IV hilar cholangiocarcinoma surgically treated in our department between Jan. 1990 and Jan. 2008 were retrospectively analyzed. Since January 2000, "Kou mode of hepatic hilar anastomosis" was performed for some patients with advanced hilar cholangiocarcinoma. The patients were divided into two groups: group A treated between 1990 and 1999, group B between 2000 and 2008. The rate of resection, therapeutic efficacy and complications in these two groups were compared, respectively., Results: Of the 37 cases with hilar cholangiocarcinoma in group A, 4 were surgically treated (10.8%), with 1 (2.7%) radical resection and 3 (8.1%) palliative resection. Among the 52 cases with hilar cholangiocarcinoma in the group B, 35 (67.3%) received surgical resection, of them 15 (28.8%) underwent radical resection and 20 (38.5%) had palliative resection. Twenty-eight of these 35 cases underwent the "Kou mode of hepatic hilar anastomosis". The resection rate of advanced hilar cholangiocarcinoma in the group B was significantly higher than that in group A (P < 0.05). The complications in the 89 cases included ascites (3 cases), hemobilia (1 case), heart failure (1 case), and wound infection (2 cases). All the patients who were treated with the "Kou mode of hepatic hilar anastomosis" developed bile leakage to a varying degree and recovered after drainage and symptomatic treatment., Conclusion: The resection rate of type III or IV advanced hilar cholangiocarcinoma can be remarkably improved by using a novel alternative surgical technique called "Kou mode of hepatic hilar anastomosis". However, the long-term outcome still needs to be determined by close follow-up and further observation.

Objective: To study the expression and distribution of hepatocyte-enriched transcriptional factors during the differentiation of hepatocyte by rat bone marrow stem cells in vitro., Methods: Mesenchymal stem cells were isolated from rat bone marrow and induced into mature hepatocyte in vitro. The mRNA expression levels of hepatocyte nuclear factor 4 alpha (HNF4 alpha), CCAAT enhancer binding protein (C/EBP) alpha and beta were compared between induced and non-induced cultures using semi-quantitative reverse transcription-polymerase chain reaction. The distribution pattern of HNF4 alpha was detected by immunofluorescence staining and observed by fluorescence microscope., Results: Transcriptional factors HNF4 alpha, C/EBP alpha, and C/EBP beta were expressed in the induced cells during the culture process. The mRNA expression levels of HNF4 alpha and C/EBP alpha were significantly higher in induced cultures than those in non-induced cultures in the early stage, whereas C/EBP beta expression was significantly up-regulated in induced cultures at the late stage (P < 0.05). Immunofluorescence staining showed that HNF4 alpha was located in the cell nucleus of differentiated cells., Conclusion: The characteristic time-dependent expression of transcriptional factors HNF4 alpha, C/EBP alpha, and C/EBP beta during the hepatocyte differentiation by bone marrow stem cells demonstrates that the expressions of these transcriptional factors are closely related to the initiation and maintenance of hepatocyte differentiation.

Objective: To analyze the clinical features of pregnancy and lactation-associated osteoporosis (PLO)., Methods: The clinical data of 4 patients with PLO were retrospectively analyzed., Results: Bone pain and limitation of motion appeared within 2 months after childbirth in these 4 patients. X-ray films showed multiple vertebral fractures of thoracic and/or lumbar spine. Three of the 4 patients had positive family history of osteoporosis or low trauma fractures. After the diagnosis of PLO was confirmed, lactation was discontinued, and treatment of calcium, active vitamin D analogues, and bisphosphonates were initiated. After these managements, no new fracture occurred and the BMD at L(2-4) was markedly increased by 12.7% - 22.7% during their follow-up period of 1 - 4 years., Conclusion: Women with positive family history of osteoporosis or low trauma fractures may be susceptible to PLO. PLO should be considered when lumbar or back pain occur during pregnancy and lactation. Besides calcium and active vitamin D analogues, anti-resorptive agents (bisphosphonate or calcitonin) may be effective on PLO.

Objective: To investigate factors that most influence urban road traffic injuries (RTI) mortality and morbidity., Methods: The study used linked police and hospital records of RTI patients in the city of Hangzhou during the 3-year period 2004-2006. Three RTI outcome groups were included: (1) fatally injured; (2) severely injured; and (3) mildly injured persons., Results: High risks for fatal road traffic accidents (RTA) were found on urban links, over weekend, during night hours, in male drivers who drove old vehicles without using seat belts, and at exceeding speeds, or with night time accidents and bad weather condition. In case of higher risk for all urban road users on urban junctions, the numbers on mildly injury cases were increasing. The highest combined risk for dying or being severely injured was found in male drivers driving at excessive speed, on urban links, and with night time accidents., Conclusions: Intensifying safety education of motor vehicle drivers, enhancing traffic management and keeping balance of "person-vehicle-road" system will greatly reduce the urban traffic accidents and casualties.

Global DNA hypomethylation in tumor tissue is a common characteristic in a variety of malignancies such as breast, colon, oral, lung, and blood cancers. A rapid and sensitive method has been developed for the determination of global DNA methylation in cells. Five substances--2'-deoxycytidine (dC), 5-methyl 2'-deoxycytidine (mdC), 2'-deoxyadenosine (dA), 2'-deoxythymidine (dT), and 2'-deoxyguanosine (dG)--were completely separated by high-performance capillary electrophoresis in 10 min. Intraday coefficient of variation was less than 1%, and interday coefficient of variation was less than 2%. The minimal detection limit was 1 microM. Acquired drug resistance to methotrexate (MTX) is one of the most serious problems in cancer chemotherapy. Under optimal conditions, we analyzed global DNA methylation levels in A549 and A549/MTX cells, and only 10(5) cells are needed to obtain reliable results. The percentage of 5-methyl-2'-deoxycytidine (5-mC) was 4.80+/-0.52% in A549 cells, and this decreased to 4.20+/-0.44% in A549/MTX cells. It was considered as statistically significant. This demonstrated that the mechanisms of acquired drug resistance to MTX might be concerned with DNA methylation.

Objective: To study the chiral selectivity in vascular endothelial differentiation in drug resistant lung cancer cells induced by high-dose L- or D-methotrexate (MTX) enantiomer., Methods: Human lung cancer cells of the line A549 were co-cultured with high-dose (15 micromol/L) L- or D-MTX enantiomer so as to develop cancer cells resistant to MTX. MTT method was used to detect the drug resistant index. Flow cytometry was used to detect the expression of CD44, a transmembrane glycoprotein reflecting the migration ability of cells, CD31, a marker of vascular endothelium, and P-170 protein. Fifteen BALB/c nude mice were inoculated with the parent A549 cells, L-MTX-resistant A549 cells induced by L-MTX enantiomer, and D-MTX-resistant A549 cells induced by D-MTX enantiomer. Four weeks later the mice were killed to take out the tumor masses. Immunohistochemistry with CD34 staining was used to detect the microvascular density (MVD)., Results: The drug resistant index of the D-MTX induced drug resistant A549 cells was 20.1 +/- 2.3, significantly higher than that of the L-MTX-induced cells (12.4 +/- 1.2, P = 0.000). The CD44 positive rate of the D-MTX induced A549 cells was 97.0% +/- 0.9%, not significantly different from that of the L-MTX-induced A549 cells (96.7% +/- 1.4%, P = 0.544). The CD31 positive rate of the D-MTX induced A549 cells was 91.9% +/- 3.2%, significantly higher than that of the L-MTX-induced A549 cells (32.9% +/- 8.0%, P = 0.000). The P-170 protein positive rate of the parent cells was 85.5% +/- 4.6%, and the P-170 protein positive rate of the D-MTX-induced A549 cells was 87.0% +/- 8.9%, significantly higher than that of the L-MTX-induced cells (71.5% +/- 8.2%, P = 0.002). The MVD of the D-MTX-induced cells was 55.9 +/- 11.9, significantly higher than that of the L-MTX-induced cells (7.2 +/- 1.7, P = 0.000). MVD was significantly positively correlated with the CD31 level (r = 0.462, P = 0.007), and not correlated with P-170 protein and CD34 levels., Conclusion: The MTX enantiomers have different chiral selectivity on human lung cancer cell, D-MTX resistant cells shows a potential of differentiation from cancer cells to vascular endothelial cells. D-MTX is not be regarded just as a pollutant in the drug MTX, MTX with single enantiomer (L-MTX) should be selected clinically so as to decrease the side effects of D-MTX.

Extracellular acidification inhibited LPS-induced TNF-alpha protein production, which was associated with an inhibition of TNF-alpha mRNA expression, in mouse peritoneal macrophages. The LPS-induced cytokine production was also inhibited by G(s) protein-coupled receptor agonists prostaglandin E(1) and isoproterenol. Among OGR1 family proton-sensing GTP-binding regulatory protein-coupled receptors, TDAG8, OGR1, and G2A are expressed in the cells. The inhibitory action by acidic pH on TNF-alpha production was significantly attenuated in macrophages from TDAG8(Tp/Tp) mice but not in those from OGR1(geo/geo) mice. Moreover, small interfering RNA specific to TDAG8, but not to G2A, clearly attenuated the acidification-induced inhibition of TNF-alpha production. On the other hand, the down-regulation or deficiency of TDAG8 hardly affected prostaglandin E(1)- or isoproterenol-induced actions. LPS-induced IL-6 production was also inhibited by extracellular acidification in a manner that was sensitive to TDAG8 expression. The acidic pH-induced inhibitory action on the cytokine production was significantly reversed either by a small interfering RNA specific to G(s) proteins or by a protein kinase A (PKA)-specific inhibitor H89. Indeed, a PKA-specific cAMP derivative inhibited LPS-induced cytokine production. Moreover, acidification induced cAMP accumulation in a TDAG8-specific way. We conclude that TDAG8, at least partly, mediates the extracellular acidification-induced inhibition of proinflammatory cytokine production through the G(s) protein/cAMP/PKA signaling pathway in mouse macrophages.

Background: Several studies in the literature have investigated the possible role of the extent of lymphadenectomy in gastric cancer treatment failure. The current study attempted to determine the effectiveness and safety of lymphadenectomy with gastrectomy for the treatment of gastric cancer., Methods: Randomized controlled trials (RCTs) were identified by means of MEDLINE, EMBASE, Cochrane Controlled Trials Register databases, and Chinese Biomedical Database, as well as by selecting references from relevant articles., Results: Overall, 14 RCTs (3,432 patients) were included in the meta-analysis. Of the D1 and D2 surgery groups, the operative mortality and postoperative morbidity were higher in the D2 group than in the D1 group, but the 3- and 5-year survival rates were not statistically different. Also the operative time was shorter in D1 compared to D2. In the D2 versus the D3 surgical group, the operative mortality, percentage of postoperative complications, operative time, and hospital stay were not significantly different., Conclusions: The results suggest that D2 and D3 surgery may not offer specific advantages for gastric cancer and instead may lead to disadvantages for patient outcomes.

Background: Numerous functional studies have shown that nitrergic neurotransmission plays a central role in peristalsis and sphincter relaxation throughout the gut and impaired nitrergic neurotransmission has been implicated in clinical disorders of all parts of the gut. However, the role of nitric oxide (NO) as a neurotransmitter continues to be controversial because: 1) the cellular site of production during neurotransmission is not well established; 2) NO may interacts with other inhibitory neurotransmitter candidates, making it difficult to understand its precise role., Methodology/principal Findings: Imaging NO can help resolve many of the controversies regarding the role of NO in nitrergic neurotransmission. Imaging of NO and its cellular site of production is now possible. NO forms quantifiable fluorescent compound with diaminofluorescein (DAF) and allows imaging of NO with good specificity and sensitivity in living cells. In this report we describe visualization and regulation of NO and calcium (Ca(2+)) in the myenteric nerve varicosities during neurotransmission using multiphoton microscopy. Our results in mice gastric muscle strips provide visual proof that NO is produced de novo in the nitrergic nerve varicosities upon nonadrenergic noncholinergic (NANC) nerve stimulation. These studies show that NO is a neurotransmitter rather than a mediator. Changes in NO production in response to various pharmacological treatments correlated well with changes in slow inhibitory junction potential of smooth muscles., Conclusions/significance: Dual imaging and electrophysiologic studies provide visual proof that during nitrergic neurotransmission NO is produced in the nerve terminals. Such studies may help define whether NO production or its signaling pathway is responsible for impaired nitrergic neurotransmission in pathological states.

Aim: To construct urothelium-specific recombinant adenovirus and investigate its inhibition in bladder cancer cell., Methods: RT-PCR analysis was used to determine expression patterns of hUPII and coxsackie adenovirus receptor on multiple cell lines. Transient transfection and luciferase detecting assay were used to detect tissue specificity of the hUPII promoter. Recombinant adenovirus Ad-UPII-E1A and Ad-UPII-Null were constructed. Restrictive enzyme digestion assay and PCR confirmed the correct construction. The adenovirus E1A protein expressed in BIU-87 was tested by Western blot after cells were infected with recombinant adenovirus. Recombinant adenovirus Ad-UPII-E1A was tested for its inhibition in bladder cancer cell line BIU-87., Results: HUPII and CAR were expressed and the hUPII promoter is highly active in bladder cancer cell line BIU-87. Using homologous recombination in bacteria technology, the hUPII promoter and E1A gene were inserted into the genome of type 5 recombinant adenovirus. The E1A protein was markedly positive in the samples of BIU-87 cells infected with recombinant adenovirus Ad-UPII-E1A. MTT assay demonstrated recombinant adenovirus Ad-UPII-E1A inhibited bladder cancer cell BIU-87 growth., Conclusion: The hUPII promoter shows high tissue specificity. Recombinant adenovirus Ad-UPII-E1A and Ad-UPII-Null were constructed and confirmed. Recombinant adenovirus Ad-UPII-E1A is effective in inhibition in bladder cancer cell line BIU-87., (Copyright 2009 S. Karger AG, Basel.)

Gastrointestinal (GI) abnormalities resulting from spinal cord injury (SCI) are challenging disorders that have not been examined experimentally using clinically relevant models. In this study, female Sprague-Dawley rats (n=5/groupx4: T10-T11 contusion, laminectomy, or naïve) were fasted for 24 h before being submitted to dye recovery assays (Phenol Red solution, 1.5 ml/rat; per oral) on GI emptying/transiting at 48 h or 4 weeks postinjury (p.i.). Compared with controls, SCI significantly increased dye recovery rate (DRR, determined by spectrophotometry) in the duodenum (+84.6%) and stomach (+32.6%), but decreased it in the jejunum (-64.1% and -49.5%) and ileum (-73.6% and -70.1%) at 48 h and 4 weeks p.i., respectively (P

LAPTM4B was proven to overexpress in hepatocellular carcinoma and relate to differentiation. We immunohistochemically investigated the expression and potential clinicopathological and prognostic significance of LAPTM4B encoded protein, LAPTM4B-35, in extrahepatic cholangiocarcinoma (EHCC) for specimens from consecutive 81 patients. LAPTM4B-35 staining was positive in cancer tissues from 59 patients (72.8%), including 12 with score 1, 22 with score 2 and 25 with score 3. No positive staining was found in non-cancer epithelia. The staining score in cancer tissues was not only significantly associated with TNM staging, histological grade, perineural and lymph node invasion (P<0.05), but also of comprehensive prognostic implications, including integrated estimation with CA19-9. These data established that LAPTM4B-35 positively expressed in a great portion of EHCC and might be a novel molecular maker of progression, invasiveness and poor prognosis.

Background: Among the deaths due to trauma, about one half of the patients suffer from road traffic injury (RTI). Most of RTI patients complicate acute respiratory distress syndrome (ARDS) and severe multiple injuries. ARDS is a major contributor to morbidity and mortality in trauma patients. Although many injuries and conditions are believed to be associated with ARDS independent risk factors in trauma patients, their relative importance in development of the syndrome are undefined. We hypothesize that not all of the traditional risk factors impacting mortality are independently associated with patients strictly identified by traffic injury. This study aimed to sieve distinctive risk factors in our RTI population, meanwhile, we also hypothesize that there may exist significantly different risk factors in these patients., Methods: This was a retrospective cohort study regarding RTI as a single cause for emergency intensive care unit (EICU) admission. Patients identified as severe RTI with post-traumatic ARDS were enrolled in a prospectively maintained database between May 2002 and April 2007 and observed. Twenty-three items of potential risk impacting mortality were calculated by univariate and multivariate Logistic analyses in order to find distinctive items in these severe RTI patients., Results: There were 247 RTI patients with post-traumatic ARDS admitted to EICU during the study period. The unadjusted odds ratio (OR) and 95% confidence intervals (CI) of mortality were associated with six risk factors out of 23: APACHE II score, duration of trauma factor, pulmonary contusion, aspiration of gastric contents, sepsis and duration of mechanical ventilation. The adjusted ORs with 95% CI were denoted with respect to surviving beyond 96 hours EICU admission (APACHE II score, duration of trauma factor, aspiration of gastric contents), APACHE II score beyond 20 EICU admission (duration of trauma factor, sepsis, duration of mechanical ventilation) and mechanical ventilation beyond 7 days EICU admission (duration of trauma factor and sepsis)., Conclusions: We have retrospectively demonstrated an adverse effect of six different risk factors out of 23 items in mortality of post-traumatic ARDS within severe RTI patients and, moreover, gained distinct outcomes in stratified patients under real emergency trauma circumstance. An impact of APACHE II score and pulmonary contusion contributing to prediction of mortality may exist in prophase after traffic injury. Sepsis is still a vital risk factor referring to systemic inflammatory response syndrome, infection, and secondary multiple organs dysfunction. Eliminating trauma factors as early as possible becomes the critical therapeutic measure. Aspiration of gastric contents could lead to incremental mortality due to severe ventilation associated pneumonia. Long-standing mechanical ventilation should be constrained on account of severe refractory complications.

Background: We carried out a meta-analysis to determine the effectiveness of breast conservation therapy (BCT) or mastectomy (MT) for stage I or stage II breast cancer., Methods: A fully recursive literature search was conducted in the Cochrane Controlled Trials Register Databases, Medline, EMBASE and Chinese Biomedical Literature Database in any language. Randomized controlled trials (RCTs) were considered for inclusion. Analyses were carried out using RevMan software., Results: In all, 18 RCTs including a total of 9388 patients were included. The meta-analysis showed that the overall survival in 3, 5, 10, 15 and 20 years and the locoregional recurrence rate in 3, 5, 15 and 20 years were not statistically significantly different between group BCT and group MT, but 10-year locoregional recurrence rate increased in group BCT. The sensitivity analysis indicated that both overall survival and locoregional recurrence rate were not statistically significant difference between group BCT and group MT. In the subgroup analysis, there was no significant difference in OS and locoregional recurrence rate between group BCT and group MT, but 20-year locoregional recurrence rate was statistically significantly higher in group BCT than group MT for women with tumors 2 cm or smaller., Conclusion: BCT was better choice than MT for women with stage I or stage II breast cancer.

Gallbladder carcinoma (GBC) is a malignancy with dismal prognosis and unclear gene expression profile. We aimed to first present the expression of LAPTM4B-35, one product of a cancer associated gene recently cloned in hepatocellular carcinoma (HCC), and its correlation with clinicopathological features and prognosis of GBC. Immunohistochemical detection of LAPTM4B-35 was performed on samples from 75 patients with GBC. LAPTM4B-35 protein was overexpressed in 57 patients (76%) with GBC. The staining scores were significantly related to histology type, lymph node involvement, distant metastasis, Nevin staging and differentiation of GBC (P<0.05). Univariate analysis revealed that overall or disease-free survival of patients was inversely associated with its staining scores (P<0.001). Multivariate analysis showed that LAPTM4B-35 staining score was an independent prognostic marker for both overall and disease-free post-resectional survival of GBC (P=0.004 and 0.027, respectively). LAPTM4B-35 overexpressed in a majority of GBCs and correlated with clinicopathological features and post-resectional survival.

Objective: To observe the viability and function of human bone marrow stem cell-derived hepatocytes following cryopreservation in vitro., Methods: Human bone marrow cells were induced to differentiate into hepatocytes in the presence of multiple factors. Mature hepatocytes were cryopreserved in 90% FBS and 10% DMSO (Group A), 10% FBS, 30% glycerol and 60% conditioned medium (Group B), and 10% FBS, 10% DMSO, and 80% UW solution (Group C). The cells were thawed after 4 weeks, and the cell viability and the albumin level were determined., Results: The human bone marrow derived hepatocytes maintained functional morphology after thawing. The viabilities in Group A, B and C were (60.0 +/- 3.3)%, (91.0 +/- 2.6)%, and (89.0 +/- 1.4)%, respectively. After culturing for 24 h, the albumin levels in Group A, B and C were (0.210 +/- 0.005) g/L, (0.340 +/- 0.020) g/L and (0.330 +/- 0.030) g/L, respectively., Conclusions: Human bone marrow stem cell-derived hepatocytes can maintain the viability and function after cryopreservation. These cells may contribute to an important source of hepatocytes for clinical hepatocyte transplantation and artificial liver support system.

A submicrometer-scaled polystyrene/melamine-formaldehyde hollow microsphere composite was prepared by self-assembling of sulfonated polystyrene (SPS) latex particles at the interface of emulsion droplets and then being fixed in place using a hard melamine-formaldehyde (MF) composite layer. For control-released purposes, the influential factors that control the size and uniformity of the packed-droplets and the permeability of the composite shell, including the initial particle location, the hydrophilicity and the size of colloidal templates, the oil phase solvent and reserving time of emulsions after the addition of MF prepolymer, were further studied. Relatively uniform sized particle packed-droplets with an average diameter of 10 microm were obtained. The assembled SPS particles kept ordering and minimal conglutination after the preparation of composite microspheres, which allows of controlling the permeability from the interstices between the particles. Porous-mesh-structured MF composite layer was formed to further control the permeability. The morphology of emulsions and composite microspheres were characterized by optical microscopy, scanning and transmission electron microscopy.

Objective: To evaluate the combined surgical therapy for calculi of intrahepatic duct., Methods: One hundred and eight cases of hepatolithiasis treated in our hospital from January 1986 to September 2003 were summarized and analyzed retrospectively. The primary surgery included 57 cases of biliary tract exploration and cholangiolithotomy, 26 cases of cholangiojejunostomy, and 25 cases of partial hepatectomy. Of these cases, 156 operations were performed totally. There were 36 patients undergoing two or more operations for cholangiolithiasis problem. Fiberoptic choledochoscopy was employed to examine and remove the residual stones during and after operation in all cases., Results: The residual stone rates were 57.89% (33/57), 26.92% (7/26), and 24.0% (6/25) for biliary tract exploration and cholangiolithotomy, cholangiojejunostomy, and partial hepatectomy, respectively. Fiberoptic choledochoscopy was utilized pre- and post-operatively for each patient, which was performed 2 to 9 times on each case. Following the combined treatment, the rate of residual stone after operation decreased to 12.96% (14/108)., Conclusions: The combined surgical therapy is valuable for resolving the problem of hepatolithiasis in most of the patients. Regarding the clinical outcomes of different surgical procedures, partial hepatectomy is superior to cholangiojejunostomy or biliary tract exploration and cholangiolithotomy. Fiberoptic choledochoscopy is also important to reduce the occurrence of residual stones and the rate of reoperation.

Objective: To observe the expression pattern of albumin during the hepatocyte differentiation by human bone marrow stem cells in vitro., Methods: Human bone marrow cells were harvested and cultured in the presence of hepatocyte growth factor (HGF), fibroblast growth factor (FGF) and lymphocyte inhibitory factor (LIF). Cells were stained immunohistochemically by albumin specific antibody and examined under a confocal microscope. Supernatant albumin level was measured biochemically on a serial time points of the culture., Results: By this condition, the attached cells became mature morphologically in 1 week of culture. Hepatocyte-specific albumin could be detected in mature cells. The albumin level revealed a time-dependent change during a 4-week culture., Conclusion: Human bone marrow cells could be induced to differentiate to mature hepatocytes that produce and secret albumin in vitro. These cells may contribute to a stable source of hepatocytes for clinical hepatocyte transplantation and artificial liver support system.

Background & Objective: Allogeneic stem cell transplantation has achieved great progress in hematology, and provided possibility to cure malignant hematological diseases. Graft-versus-host disease (GVHD) is a significant complication after transplantation with a mortality of over 30%. In clinical practice we noticed that the successful implantation and the severity of GVHD were associated with T cell ratio of graft to receptor after pretreatment. This study was to set up rat allo-stem cell transplantation model, and explore effects of different T cell ratios of graft to receptor on severity of GVHD., Methods: SD rats were selected as receptor, and pretreated with total body irradiation (TBI), and cyclophosphamide (CTX). WISTAR rats were used as donors, their bone marrow cells were collected as graft. Receptors were transfused with different ratios of T cells (1:1, 2:1, and 4:1). Life-span, clinical and pathologic GVHD scores were observed., Results: When T cell ratios of graft to receptor were 1:1, 2:1, and 4:1, life-spans of receptors were (16.7+/-1.3) d, (16.8+/-1.5) d, and (15.0+/-1.7) d, respectively; clinical GVHD scores of receptors were 2.4+/-0.6, 3.3+/-1.2, and 5.3+/-1.4, respectively. Life-span, and clinical and pathologic GVHD scores of receptors transfused with T cells of 1:1 and 2:1 were significantly different from those of receptors transfused with T cells of 4:1(P<0.05)., Conclusion: When T cell ratio of graft to receptor was 1:1, or 2:1, the GVHD was comparatively mild; when this ratio reached to 4:1, the GVHD was aggravated.

Objective: To evaluate the diagnosis and treatment of mesenteric venous thrombosis., Methods: The clinical data of 11 cases diagnosed as mesenteric venous thrombosis between 1992 and 2001 in PUMC Hospital were analyzed retrospectively., Results: Postoperative state(27.3%), especially cirrhosis and portal hypertension, and other history of thrombosis (27.3%) were the most common causes. Thrombolysis was performed successfully in two of the eleven cases. The rest of them were misdiagnosed in other hospitals and operated. No patient died after operation, and one (11.1%) recurrence was found., Conclusions: Early application of anticoagulant is necessary for patients with thrombosis risks. For suspected patients, early computed tomography (CT) and DSA examination should be performed, and prompt thrombolysis and anticoagulation therapy can be performed to avoid the bowel resection after definite diagnosis. To reduce the recurrence, anticoagulant should be maintained for a proper time.

Objective: To evaluate the tolerance limit of rats to normothermic hepatic inflow occlusion under portal blood bypass., Methods: A new rat model of normothermic hepatic inflow occlusion under portal blood bypass was established by clamping temporarily the pedicles of all liver lobes while the caudal lobe was kept as a passage of the portal blood flow. After hepatic blood flow restored, the caudal lobe was cut off. On the 7th postoperative day, survival rate, hepatic morphological changes, and the severity and reversibility of the injured energy metabolism of the liver were investigated., Results: All rats that had been subjected to 30, 60 and 90 minutes of hepatic inflow occlusion under portal blood bypass survived on the 7th postoperative day. ischemia-reperfusion injury of the liver was reversible and compensatory in rats with hepatic inflow occlusion within 90 minutes. However, the survival rates of rats with 100, 110 and 120 minutes of hepatic inflow occlusion were 50%, 30% and 20% respectively. Liver injury of rats with 120 minutes of hepatic inflow occlusion was severe and Irreversible., Conclusions: The tolerance limit of rats to normothermic hepatic inflow occlusion is enhanced significantly under portal blood bypass and the upper limit is 90 minutes.

An inwardly rectifying K(+) conductance closely resembling the human ether-a-go-go-related gene (HERG) current was identified in single smooth muscle cells of opossum esophageal circular muscle. When cells were voltage clamped at 0 mV, in isotonic K(+) solution (140 mM), step hyperpolarizations to -120 mV in 10-mV increments resulted in large inward currents that activated rapidly and then declined slowly (inactivated) during the test pulse in a time- and voltage- dependent fashion. The HERG K(+) channel blockers E-4031 (1 microM), cisapride (1 microM), and La(3+) (100 microM) strongly inhibited these currents as did millimolar concentrations of Ba(2+). Immunoflourescence staining with anti-HERG antibody in single cells resulted in punctate staining at the sarcolemma. At membrane potentials near the resting membrane potential (-50 to -70 mV), this K(+) conductance did not inactivate completely. In conventional microelectrode recordings, both E-4031 and cisapride depolarized tissue strips by 10 mV and also induced phasic contractions. In combination, these results provide direct experimental evidence for expression of HERG-like K(+) currents in gastrointestinal smooth muscle cells and suggest that HERG plays an important role in modulating the resting membrane potential.

Background & Aims: Mesenteric ischemia, infection, and inflammatory bowel disease may eventuate in severe colitis, complicated by toxic megacolon with impending intestinal perforation. Monochloramine (NH(2)Cl) is a membrane-permeant oxidant generated during colitis by the large amount of ambient luminal NH(3) in the colon. Reactive oxygen metabolites can modulate smooth muscle ion channels and thereby affect colonic motility, which is markedly impaired in colitis., Methods: Effects of NH(2)Cl on ionic currents in the innermost smooth muscle layer of the colon, the tunica muscularis mucosae, were examined using the patch clamp technique. Membrane potential in whole tissue strips was measured using high-resistance microelectrodes., Results: Whole cell voltage clamp experiments showed that NH(2)Cl (3-30 micromol/L) enhanced outward currents in a dose-dependent manner, increasing currents more than 8-fold at a test potential of +30 mV. Tail current analysis showed that the currents enhanced by NH(2)Cl were K(+) currents. Inhibition by tetraethylammonium and iberiotoxin suggested that these currents represented activation of large-conductance, Ca(2+)-activated K(+) channels. The membrane-impermeant oxidant taurine monochloramine, however, had no effect on whole cell currents. Single-channel studies in inside-out patches showed that NH(2)Cl increased open probability of a 257-pS channel in symmetrical (140 mmol/L) K(+). In the presence of NH(2)Cl, the steady-state voltage dependence of activation was shifted by -22 mV to the left with no change in the single-channel amplitude. The sulfhydryl alkylating agent N-ethylmaleimide prevented NH(2)Cl-induced channel activation. NH(2)Cl also hyperpolarized intact muscle strips, an effect blocked by iberiotoxin., Conclusions: NH(2)Cl, at concentrations expected to be found during colitis, may contribute to smooth muscle dysfunction by a direct oxidant effect on maxi K(+) channels.

We investigated the effects of the sensory neuropeptide substance P (SP) on amylase and fluid secretion in the isolated vascularly perfused rat pancreas. SP inhibited CCK-induced amylase release and secretin-induced juice flow via the pancreatic duct in a dose-related fashion. Threshold inhibition occurred following addition of 10(-10) M SP to the perfusate, and maximal inhibition was seen with 10(-8) M SP. The effects of SP were partially blocked by both the neurokinin-1 (NK1) and neurokinin-2 (NK2) receptor antagonists. Atropine and TTX blocked SP-induced effects on both amylase secretion (26 and 63% blockade, respectively) and pancreatic juice flow (21 and 79% blockade, respectively). Excitation of pancreatic sensory nerves using capsaicin (in the absence of SP) inhibited both amylase and pancreatic juice flow via activation of the NK1 receptor. We conclude that SP inhibits exocrine secretion via an indirect neural mechanism.

A nitric oxide (NO)-like product of the L-arginine NO synthase pathway has been shown to be a major inhibitory neurotransmitter that is involved in the slow component of the inhibitory junction potential (IJP) elicited by stimulation of nonadrenergic, noncholinergic nerves. However, the exact nature of the nitrergic transmitter, the role of cGMP, and the involvement of a potassium or a chloride conductance in the slow IJP remain unresolved. We examined the effects of soluble guanylate cyclase inhibitors LY-83583 and 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ), potassium-channel blockers and putative chloride-channel blockers diphenylamine-2-carboxylate (DPC) and niflumic acid (NFA) on the hyperpolarization elicited by an NO. donor, diethylenetriamine/NO adduct (DNO), NO in solution, and an NO+ donor, sodium nitroprusside (SNP), in the guinea pig ileal circular muscle. Effects of these blockers on purinergic (fast) and nitrergic (slow) IJP were also examined. DNO-induced hyperpolarization and nitrergic slow IJP were suppressed by LY-83583 or ODQ and DPC or NFA but not by the potassium-channel blocker apamin. In contrast, hyperpolarization caused by SNP or solubilized NO gas and purinergic fast IJP were antagonized by apamin but not by inhibitors of guanylate cyclase or chloride channels. These results demonstrate biological differences in the actions of different redox states of NO and suggest that NO. is the nitrergic inhibitory neurotransmitter.

Unlabelled: Mice lacking neuronal nitric oxide synthase gene (ncNOS) were used to determine the enzymatic source of nitric oxide (NO) and its relationship with other putative inhibitory neurotransmitters. Inhibitory junction potentials (IJP) of circular smooth muscle of gastric fundus were studied. The IJP in the wild-type mice consists of overlapping components, the fast and slow IJPs. NOS inhibitor L-NA or VIP receptor antagonist VIP(10-28), blocks the slow IJP but not the fast IJP. The fast UP is blocked by alpha-beta methylene ATP tachyphylaxis, by reactive blue 2, and by apamin. The IJP in the ncNOS-deficient [ncNOS(-)] mutant is of short duration and is abolished by blockers of the fast IJP, but is unaffected by blockers of the slow UP. Exogenous VIP produces membrane hyperpolarization in strips from wild-type but not ncNOS(-) mice. The hyperpolarizing action of VIP is resistant to nifedipine but is sensitive to omega-conotoxin GVIA., In Conclusion: (a) NO derived from ncNOS is an inhibitory neurotransmitter rather than a postjunctional mediator; (b) VIP is a prejunctional neurotransmitter that causes release of evanescent NO; and (c) ATP acts in parallel with the VIP/NO pathway.

The Influence of energy transfer inhibitors of ATP synthase on the chlorophyll fluorescence quenching of thylakoids was studied by the saturation pulse method. Triphenyltin chloride (TPT) treatment could result in an increase of q(Q) and a decrease of q(E) of the thylakoids, but DCCD could not. This increase could be abolished when the deltapH across the thylakoid membrane was dissipated by uncouplers (1O mM NH(4)Cl plus 1&mgr;M nigericin) or in higher salt medium which could relieve localized protons on the thylakoid membrane. In the electron transport system of H(2)O right curved arrow PD(0X) or H(2)O right curved arrow PBQ coupled with PSII. The stimulatory effect of TPT on q(Q) also diminished. By the methods of modulated fluorescence and low temperature fluorescence measurement we observed that TPT markedly increased the imbalance of the distribution of light energy between PSI and PSII in favor of PSII. These results suggest that the CF(0) proton flow could influence the light energy distribution between the two photosystems. This might be materialized by the regulation of the membrane-localized protons on the photochemical efficiency of photosystem II.

The article describes the effectiveness of compatible application of heat-clearing and detoxifying drugs with blood circulation improving drugs to the animal model with endotoxemia and nonspecific inflammation. The compatible application reduces PGE2, endotoxin blood concentration and reduced viscosity of whole blood, decreases Evans blue extravasation volume and pes swelling percentage, increases serum cortisol content and enhances fibrinolytic activity. The experimental result shows that in most cases these two drugs work better when used in combination which implies that compatible application is more effective in detoxification, antiinflammation and inflammation recovery.

1. Intracellular membrane potential recordings were made from circular smooth muscle cells of the guinea-pig ileum in the presence of atropine (1 microM) and nifedipine (0.1 microM) at 30 degrees C. 2. Electrical field stimulation with one or four pulses produced a fast inhibitory junction potential (IJP) which lasted around 1 s. It was abolished by tetrodotoxin (1 microM), apamin (0.3 microM), and alpha, beta-methylene ATP tachyphylaxis. 3. Nitric oxide synthase inhibitor N-nitro-L-arginine (L-NNA; 200 microM) had no effect on the resting membrane potential or the fast IJP. 4. Electrical field stimulation in the presence of apamin and substance P desensitization produced a slow IJP which was abolished by tetrodotoxin (1 microM). 5. L-NNA significantly reduced the amplitude of the slow IJP (P < 0.01). The antagonistic effect of L-NNA was reversed by L-arginine but not by D-arginine. 6. Injections of alpha, beta-methylene ATP, nitric oxide (NO), and vasoactive intestinal polypeptide (VIP) into the recording chamber caused tetrodotoxin-resistant hyperpolarizations of the smooth muscle membrane. Substance P desensitization did not modify the amplitudes of the hyperpolarizing response to ATP or NO, but increased the VIP hyperpolarization by 150% (P < 0.01). 7. L-NNA did not modify the amplitude of hyperpolarization due to ATP or NO; however, it antagonized VIP-induced hyperpolarization (P < 0.01). 8. These studies show that in the guinea-pig ileum circular muscle: (a) NO is not involved in the fast IJP which is mediated by ATP; (b) NO is involved in the slow IJP which is mediated by VIP and NO acting in series, and (c) the hyperpolarizing effects of VIP and the slow IJP are normally masked by overlapping depolarization due to concomitant release of substance P by the peptide VIP.

1. Intracellular membrane potential recordings were made from circular smooth muscle cells of the guinea-pig ileum in the presence of atropine (1 microM) and nifedipine (0.1 microM) at 30 degrees C. 2. Perfusion with adenosine triphospate (ATP, 100 microM) and vasoactive intestinal peptide (VIP, 2 microM) resulted in membrane hyperpolarizations of 6.4 +/- 0.3 and 6.8 +/- 0.3 mV, respectively. Picospritzes of ATP (10 mM in pipette) and VIP (100 microM in pipette) resulted in membrane hyperpolarizations of 6.9 +/- 0.4 and 6.3 +/- 0.4 mV, respectively. 3. The ATP-induced hyperpolarizations were antagonized by alpha, beta-methylene ATP desensitization (100 microM for 30 min) and the ATP antagonist Reactive Blue 2 (200 microM), but were unaffected by the VIP antagonist VIP 10-28 (1 microM). 4. The VIP-induced hyperpolarizations were antagonized by VIP 10-28, but unaffected by alpha, beta-methylene ATP desensitization and Reactive Blue 2. 5. A single pulse of transmural nerve stimulation (2 ms, 15 mA) resulted in an inhibitory junction potential (IJP) that reached a maximal amplitude of 12.9 +/- 0.5 mV at 378 +/- 20 ms from the stimulus. This fast IJP was abolished by apamin (2 microM) or tetrodotoxin (1 microM), antagonized by alpha, beta-methylene ATP desensitization or Reactive Blue 2, but unaffected by VIP 10-28. 6. In the presence of apamin (1 microM), four pulses of transmural stimulation (2 ms, 20 Hz, 15 mA) resulted in an IJP that reached a maximal amplitude of 4.8 +/- 0.2 mV at 1.4 +/- 0.1 s from the stimulus. This slow IJP was antagonized by tetrodotoxin (1 microM) or VIP 10-28 (1 microM), augmented by Reactive Blue 2 (200 microM), and unaffected by alpha, beta-methylene ATP desensitization. 7. These findings provide evidence that both ATP and VIP are inhibitory neurotransmitters in the circular muscle layer of the ileum and that ATP may be the neurotransmitter responsible for the fast IJP and VIP the neurotransmitter responsible for the slow IJP.

Intracellular recordings were made from circular smooth muscle cells of the opossum esophagus. Inhibitory junction potentials (IJPs) 8.3 +/- 0.7 mV in amplitude were observed in response to a single pulse (1 ms, 15 mA) of transmural nerve stimulation. Potassium channel blockers apamin (1 microM), tetraethylammonium (10 mM), 4-aminopyridine (250 microM), and cesium chloride (10 mM) did not reduce IJP amplitude. Conditioning hyperpolarizations to the equilibrium potential for potassium were associated with a significant increase in IJP amplitude. A small increase in membrane resistance was observed during the IJP. Changes in external potassium concentration had no significant effect on IJP amplitude acutely. However, prolonged perfusion with potassium-free Krebs solution resulted in a marked decrease in IJP amplitude as did prolonged perfusion with ouabain (0.1 mM). Low-chloride solution (12.4 mM) resulted acutely in an increase in IJP amplitude. Prolonged low-chloride perfusion resulted in a significant decrease in IJP amplitude. The anion exchange chloride channel inhibitor 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (600 microM) significantly reduced IJP amplitude. These findings suggest that the IJP observed in opossum esophageal circular smooth muscle in response to a single pulse of stimulation is due to a decrease in membrane chloride conductance. The ability of prolonged application of Na-K pump inhibitors to abolish the IJP appears to be due to known secondary effects of these agents in depleting intracellular chloride.

Junction potentials were recorded from circular muscle cells of the guinea pig ileum at various sites oral and anal to a transmural stimulus in the presence of atropine, apamin, and substance P antagonism (desensitization) at 30 degrees C. A short train of pulses produced an inhibitory junction potential (slow IJP), which preceded an excitatory junction potential (slow EJP). The slow IJP was observed up to 56.4 +/- 2.9 mm oral and 65.4 +/- 2.2 mm anal to the stimulus. The slow EJP was observed up to 50.4 +/- 1.9 mm oral and 58.3 +/- 2.1 mm anal to the stimulus. Hexamethonium (400 microM) decreased the amplitudes of the slow IJP and slow EJP at all sites. After hexamethonium, the slow IJP was observed up to 37.3 +/- 2.3 mm oral and 39.8 +/- 2.1 mm anal to the stimulus and the slow EJP was 44.2 +/- 2.5 mm oral and 43.3 +/- 2.6 mm anal to the stimulus. The slow IJP and slow EJP were associated with an increase and decrease in membrane resistance, respectively. Conditioning depolarizations of the circular muscle cells reduced the amplitudes of the slow IJP and slow EJP. Both were abolished at a membrane potential of approximately -25 mV. Conditioning hyperpolarizations increased the amplitude of both the slow IJP and slow EJP. Ionic substitution experiments with low external chloride solution (12.4 mM) resulted in an immediate increase in slow IJP and slow EJP amplitudes, whereas more prolonged perfusion resulted in a significant decrease in slow IJP and slow EJP amplitudes. 4,4'-Diisothiocyanostilbene-2,2'-disulfonic acid (400 microM) resulted in decreases in slow IJP and slow EJP amplitudes. These results suggest that the slow IJP and slow EJP are due to a decrease and increase in membrane chloride conductance, respectively. The noncholinergic neural pathways responsible for the slow IJP and slow EJP extend approximately 40 mm orally and anally along the longitudinal axis of the guinea pig ileum.

The effect of substance P antagonism on membrane potential responses to transmural nerve stimulation in the presence of atropine was examined in circular smooth muscle of the guinea pig ileum. Intracellular recordings of membrane potential responses recorded 3-5 mm oral to the transmural stimulus consisted of an inhibitory junction potential followed by two distinct depolarizations referred to as early and late excitatory junction potentials. Substance P antagonism was achieved by desensitization with high doses of substance P or use of the antagonist Spantide (Sigma Chemical Co., St. Louis, MO). Substance P antagonism had no effect on the amplitude of the inhibitory junction potential, caused an increase in the latter portion of the early excitatory junction potential, and abolished the late excitatory junction potential. The excitatory junction potential potentiated by substance P receptor antagonism was associated with a decrease in membrane resistance, increased in amplitude with conditioning hyperpolarizations to the estimated equilibrium potential for K+, and was blocked by the Cl-/HCO3- exchange inhibitor DIDS or prolonged perfusion with low-chloride solution. These studies suggest that a noncholinergic, non-substance P neurotransmitter is released from enteric motoneurons that produces excitation through an increase in smooth muscle chloride conductance.

We sought to characterize the projections of nonmuscarinic intramural inhibitory and excitatory neural input to the circular muscle layer of the small intestine. Intracellular membrane potential recordings were made in smooth muscle cells from the distal ileum of 22 guinea pigs. Intramural nerve stimulation (4 pulses, 15 mA, 20 Hz, 2 ms) was delivered by two electrodes located at varying distances from the recording electrode along the longitudinal axis of the intestine. The membrane potential responses immediately oral to the stimulus consisted of an abrupt-onset inhibitory junction potential (IJP) followed by two distinct depolarizations, an early excitatory junction potential (EJP) and a late EJP. The early IJP and early and late EJPs were observed as far as 53 +/- 5, 56 +/- 6, and 39 +/- 5 mm oral to the stimulus, respectively. The responses immediately anal to the stimulus consisted of an initial IJP followed by a prolonged hyperpolarization, which was sometimes interrupted by a superimposed EJP. The early IJP, EJP, and late IJP were observed as far as 77 +/- 12, 66 +/- 10, and 37 +/- 13 mm anal to the stimulus, respectively. All excitatory events greater than 5-10 mm oral or anal to the stimulus were blocked by hexamethonium. Hexamethonium slightly antagonized the early IJP oral to the stimulus but had no effect on the early IJP anal to the stimulus. The late IJP was not affected by hexamethonium. These studies are the first to describe a late IJP, which is observed only at sites anal to the stimulus, and a late EJP, which is observed only at sites oral to the stimulus. The neural pathways involved in these late synaptic events (late EJP and IJP) as well as the previously described early synaptic events (early EJP and IJP) are discussed.