Presentation of oropharyngeal dysphagia and rehabilitative intervention following esophagectomy: a systematic review.

No study has systematically reviewed the evidence on presentation of oropharyngeal dysphagia and swallowing rehabilitation following esophagectomy. The purposes of this systematic review are to 1) qualitatively synthesize the current findings on oropharyngeal swallowing abnormalities identified by instrumental swallowing evaluations, 2) describe the reported health-related outcomes in relation to swallowing abnormality following esophagectomy, and 3) examine the efficacy of reported rehabilitative interventions for oropharyngeal dysphagia in patients who underwent esophagectomy. Publications were searched using five electronic databases. No language or publication date restrictions were imposed. Two authors performed a blind review for published or unpublished studies that reported swallowing biomechanics and dysphagic symptoms using instrumental evaluation of swallowing, specifically the videofluoroscopic swallowing study and fiberoptic endoscopic evaluation of swallowing, and/or health-related outcomes in relation to swallowing abnormalities, and/or therapeutic interventions for oropharyngeal dysphagia following esophagectomy. Twelve studies out of 2,193 studies including 458 patients met the inclusion criteria. Reported abnormal swallowing biomechanics included vocal fold immobility, delayed onset of swallowing, reduced hyolaryngeal elevation, and reduced opening of the upper esophageal sphincter. Aspiration (0-81%) and pharyngeal residue (22-100%) were prevalent. Those abnormal swallowing biomechanics and swallowing symptoms were commonly reported following both transhiatal and transthoracic esophagectomy. Pneumonia presented in 5-25% of the study patients. One quasi-experimental study examined the effectiveness of swallowing exercises for postoperative oropharyngeal dysphagia; three case series reported a benefit of the chin-tuck maneuver in reducing aspiration and residue. This review revealed distinct swallowing impairments and increased pneumonia risks following esophagectomy. This review also found that evidence on the efficacy of therapeutic interventions was limited. Future studies are warranted to develop effective rehabilitative interventions for postesophagectomy patients with oropharyngeal dysphagia.

Effect of rehabilitation on mortality of patients with Guillain-Barre Syndrome: a propensity-matched analysis using nationwide database.

BACKGROUND: Rehabilitation for patients with Guillain-Barre Syndrome (GBS) is recommended as it improves the outcome of neurological deficits. Few studies focused on the effect of rehabilitation on mortality of the patients. AIM: To investigate the effect of rehabilitation on hospital mortality of patients with GBS using the Japanese Diagnosis Procedure Combination (DPC) nationwide administrative claims database. DESIGN: A retrospective observational cohort study. SETTING: Hospitals adopting the Japanese DPC system. POPULATION: Patients hospitalized with a diagnosis of GBS between July 2007 and October 2011. METHODS: Data analyzed included sex, age, Barthel index at admission, use of ventilation, immune therapy, and rehabilitation during hospitalization, comorbidity, hospital volume, type of hospital, and in-hospital death. One-to-one propensity score-matching was used to compare hospital mortality rates within 30- and 90-days after admission in rehabilitation and non-rehabilitation groups. The adjusted odds ratios of rehabilitation to hospital mortality were also estimated. RESULTS: A total of 3835 patients were identified and analyzed. Patients with advancing age, lower Barthel index at admission, comorbidities, ventilation, or immune therapy were more likely to receive rehabilitation during hospitalization. Propensity-matched analysis of 926 pairs showed that the rehabilitation group had lower hospital mortality rates within both 30- and 90-days than the non-rehabilitation group. The adjusted odds ratios of rehabilitation to hospital mortality within 30- and 90-days were 0.14 and 0.23, respectively. CONCLUSION: After matching patients' background, rehabilitation was associated with lower hospital mortality of patients with GBS. CLINICAL REHABILITATION IMPACT: Rehabilitation treatment is essential for patients with GBS to improve their survival.

Endoscopic treatment for early stage colorectal tumors: the comparison between EMR with small incision, simplified ESD, and ESD using the standard flush knife and the ball tipped flush knife.

BACKGROUND: Early stage colorectal tumors can be removed by endoscopic mucosal resection but larger such tumors (20 mm) may require piecemeal resection. Endoscopic submucosal dissection (ESD) using newly developed endo-knives has enabled en-block resection of lesions regardless of size and shape. However ESD for colorectal tumor is technically difficult. Therefore, we performed EMR with small incision (EMR with SI) for more reliable EMR, ESD with snaring (simplified ESD) and ESD using the standard Flush knife and the novel ball tipped Flush knife (Flush knife BT) for easier and safer colorectal ESD. AIMS: The aims of our study were (1) to compare the treatment results of the following 3 methods (EMR with SI/si-mplified ESD/ESD) for early stage colorectal tumors, and (2) to assess the performance of Flush knife BT in colorectal ESD. METHODS: We treated 24/44/468 colorectal tumors and examined the clinicopathological features and treatment results such as tumor size, resected specimen size, procedure time, en-bloc resection rate, complication rate. We also treated 58 colorectal tumors (LST-NG:20, LST-G:36, other:2) using standard Flush knife and 80 colorectal tumors (LST-NG:32, LSTG:44, other:2) using Flush knife BT, and examined the clinicopathological features and treatment results mentioned above and also the procedure speed. RESULT: The median tumor size (mm) (EMR with SI/ simplified EMR/ESD) was 20/17/30 (EMR with SI vs. simplified ESD: p = n.s, simplified ESD vs. ESD: p < 0.0001). The median resected specimen size (mm) was 22.5/26/41 (EMR with SI vs. simplified ESD: p = 0.0018, simplified ESD vs. ESD: p < 0.0001). The procedure time (min.) was 19/27/60 (EMR with SI vs. simplified ESD: p = n.s, simplified ESD vs. ESD: p < 0.0001) The en-block resection rate (%) was 83.3/90.9 /98.9. The complication rate (post-operative bleeding rate/perforation p=n.s). In the treatment results of ESD for LSTs by knives, there was no difference between standard Flush knife and Flush knife BT for clinicopathological features and treatment results (procedure time, complication rate and en bloc R0 resection rate). However, procedure speed (cm2/min.) of LST-G was significantly faster in the Flush knife BT than in standard Flush knife. (standard Flush knife: 0.21 vs. Flush knife BT: 0.27, p = 0.034). CONCLUSION: EMR with small incision (EMR with SI) and ESD with snaring (simplified ESD) are good option to fill the gap between EMR and ESD in the colorectum, and also considered to become the nice training for the introduction of ESD. Flush knife BT appears to improve procedure speed compared with standard Flush knife, especially for LST-G in colo-rectal ESD.

The performance of a novel ball-tipped Flush knife for endoscopic submucosal dissection: a case-control study.

BACKGROUND: Endoscopic submucosal dissection (ESD) using short needle knives is safe and effective, but bleeding is a problem due to low haemostatic capability. AIM: To assess the performance of a novel ball-tipped needle knife (Flush knife-BT) for ESD with particular emphasis on haemostasis. METHODS: A case-control study to compare the performance for ESD of 30 pairs of consecutive early gastrointestinal lesions (oesophagus: 12, stomach: 32, colorectum: 16) with standard Flush knife (F) vs. Flush knife-BT (BT). Primary outcome was efficacy of intraprocedure haemostasis. Secondary outcomes included procedure time, procedure speed (dividing procedure time into the area of resected specimen), en bloc resection rate and recurrence rate. RESULTS: Median intraoperative bleeding points and bleeding points requiring haemostatic forceps were smaller in the BT group than in the F group (4 vs. 8, P < 0.0001, 0 vs. 3, P < 0.0001). There was no difference between groups for procedure time; however, procedure speed was shorter in the BT group (P = 0.0078). En bloc and en bloc R0 resection rates were 100%, with no perforation or post-operative bleeding. No recurrence was observed in either group at follow-up 1 year postprocedure. CONCLUSIONS: Ball-tipped Flush knife (Flush knife-BT) appears to improve haemostatic efficacy and dissection speed compared with standard Flush knife.

Retrospective study of technical aspects and complications of endoscopic submucosal dissection for laterally spreading tumors of the colorectum.

BACKGROUND AND STUDY AIMS: Laterally spreading tumors - non granular type (LST-NG) are more often considered candidates for endoscopic submucosal dissection (ESD) than laterally spreading tumors - granular type (LST-G), because of their higher potential for submucosal invasion. However, ESD for LST-NG can be technically difficult. The aim of our study was to compare our ESD results for LST-NG and for LST-G. PATIENTS AND METHODS: Ninety-nine LST-NG and 169 LST-G measuring 20 mm in size or more were removed by ESD. We retrospectively evaluated the clinicopathological features of the tumors and treatment results (en bloc resection rate, procedure time and speed, rate of use of ancillary devices, and complication and recurrence rates). RESULTS: Histopathology revealed that there were more submucosally invasive lesions in the LST-NG than in the LST-G group (28 % vs. 9 %; P < 0.0001). The en bloc resection rate, en bloc R0 resection rate, and en bloc curative resection rate of LST-NG were similar to those of LST-G (LST-NG: 99 %, 98 %, and 88 %; LST-G: 99 %, 98 %, and 91 %). In LST-NG, the median procedure time tended to be longer (LST-NG: 69 min; LST-G: 60 min) and the median procedure speed was slower (LST-NG: 0.15 cm (2)/min; LST-G: 0.25 cm (2)/min; P < 0.0001). Use of ancillary devices was higher for LST-NG (38 % vs. 15 % for LST-G; P < 0.0001), as was the perforation rate (5.1 % vs. 0.6 % for LST-G; P = 0.027). No recurrence was seen in either group. CONCLUSIONS: ESD was an effective treatment method for both LST-NG and LST-G. However, the degree of technical difficulty appears higher for LST-NG than for LST-G lesions, as shown by the lower dissection speed and higher perforation rate. ESD for LST-NG should probably be performed by those with significant experience of colorectal ESD.

A comparison of radiation treatment plans using IMRT with helical tomotherapy and 3D conformal radiotherapy for nasal natural killer/T-cell lymphoma.

The purpose of this study was to compare radiation treatment plans (RTPs) that used intensity-modulated radiation therapy (IMRT) with helical tomotherapy (HT) or three-dimensional conformal radiation therapy (3D-CRT) for nasal natural killer/T-cell lymphoma (NNKTL). We created RTPs that used IMRT with HT or 3D-CRT for eight NNKTL patients previously treated at our institution and conducted a pilot comparison between the two modalities using the parameters of the target coverage and homogeneity for the planning target volume (PTV) and the maximum and mean doses for organs at risk (OARs). The clinical target volume (CTV) included the gross tumour volume with an additional margin of 1.5 cm and the nasopharynx, palates and nasal cavity; the PTV with the CTV plus a 2 mm margin received a total dose of 50 Gy. IMRT achieved significantly better PTV coverage, with more than 99% of the PTV receiving 90% and 95% of the prescribed dose, whereas 3D-CRT could not provide adequate coverage of the PTV, with 89.1+/-2.6% and 84.5+/-2.7% of the PTV receiving 90% and 95% of the prescribed dose, respectively (both p <0.0001). The homogeneity index was 0.29+/-0.06 for IMRT and 0.046+/-0.022 for 3D-CRT, which was statistically significant (p <0.0001). IMRT tended to provide equivalent or slightly better OAR avoidance than 3D-CRT. In conclusion, 3D-CRT could not provide adequate coverage of the PTV because the PTV was close to many OARs. IMRT should be used for NNKTL because a lack of optimal RTPs could cause local failure.

Helical tomotherapy for brain metastases: dosimetric evaluation of treatment plans and early clinical results.

The purpose of this study was to evaluate the feasibility and treatment plans of intensity-modulated radiation therapy using helical tomotherapy (HT) for brain metastases. Twenty-three patients with 1 to 4 brain metastases were treated with HT. In combination with whole-brain radiotherapy (simultaneous plans), metastatic lesions, and the whole brain were treated with 50 Gy and 30 Gy, respectively, in 10 fractions, with a simultaneous integrated boost technique. In patients treated for brain metastases alone (focal plans), metastatic lesions were treated with 35 or 37.5 Gy in 5 fractions. The treatment plans were compared regarding the conformation number (CN) and homogeneity index (HI), and differences in these indexes between simultaneous and focal plans were examined by Student's t-test. Seven and 16 patients were treated with simultaneous plans and focal plans, respectively. The mean +/- SD of CN and HI values were 0.75 +/- 0.13 and 0.063 +/- 0.042, respectively, for simultaneous plans, and 0.73 +/- 0.12 and 0.052 +/- 0.023, respectively, for focal plans. The CN and HI between the two plans were not significantly different. Response rates in 13 patients with follow-up imaging were approximately 90% for both plans and the local control rate at 1 year was 69%. One patient with a huge tumor (34.0 cc) and WHO performance status 3 treated with focal plans experienced severe headache, requiring prolongation of the treatment time, and died at 8 days after completion of treatment. The exact cause of deterioration was uncertain as no radiological investigation was performed in this patient. No late complications were observed during follow-up periods up to 20 months. HT is a viable non-invasive technique for treatment of brain metastases and achieves high accuracy in terms of dose conformity and homogeneity.

The results and limitations of endoscopic submucosal dissection for colorectal tumors.

In the colorectal tumor, the lesions suitable for the endoscopic treatment are those with no lymph node metastasis such as adenomas, intramucosal cancers, and minimally invasive submucosal cancer (invasion depth 1000 m, well and moderately differentiated type, no lymphovascular invasion). The new endOscopic technique, endoscopic submucosal dissection (ESD) enables en-bloc resection of the lesions regardless of their size and location. In order to perform ESD more easily, safely, and efficiently, we invented water jet short needle knives (Flush knife). Emitting a jet of water from the tip of a sheath enables submucosal local injection with a knife itself without replacement of operative instruments, which leads to efficient treatment. Especially, Flush knife is very effective for the lesions located at lower rectum and anal canal where there are many vessels. We treated a total of 361 colorectal lesions by ESD between June 2002 and July 2007, and en-block complete resection rate was 98.3 %. In 12 cases, "muscle retracting sign" was recognized. This sign is an index of the discontinuation of ESD, but it is impossible to diagnose preoperatively. The postoperative bleeding occurred in 0.8 % (3 cases: no blood transfusion is needed). The intraoperative perforation occurred in 1.9 % (6 cases: 5 cases were treated conservatively, 1 case was treated surgically) and the postoperative perforation occurred in 1 case (0.3%) treated surgically. ESD is the extremely effective treatment for the colorectal tumors and also is possible to be performed safely with the appropriate choice of the devices and strategy for dissection.

Intra-arterial chemoradiotherapy for locally advanced oral cavity cancer: analysis of therapeutic results in 134 cases.

The objective of this study was to investigate the therapeutic results of arterial injection therapy via the superficial temporal artery for 134 cases of stages III and IV (M0) oral cavity cancer retrospectively, and to clarify the prognostic factors. We administered intra-arterial chemoradiotherapy by continuous infusion of carboplatin in 65 cases from January 1993 to July 2002. Systemic chemotherapy was performed on 26 cases at the same time. We administered intra-arterial chemoradiotherapy by cisplatin with sodium thiosulphate in 69 cases from October 2002 to December 2006. Systemic chemotherapy was performed on 48 cases at the same time. The 3-year local control rate was 68.6% (T2-3: 77.9%; T4: 51.3%), and the 3-year survival rate was 53.9% (stage III: 62.9%; stage IV: 45.3%). Regarding the results of multivariate analysis of survival rates, age (<65), selective intra-arterial infusion, and the use of cisplatin as an agent for intra-arterial infusion were significant factors. The therapeutic results of intra-arterial chemoradiotherapy via the superficial temporal artery were not inferior to the results of surgery. In particular, the results of arterial injection therapy by cisplatin with sodium thiosulphate were excellent, so we believe that it will be a new therapy for advanced oral cavity cancer.

Excitatory GABAergic synaptic potentials in the mesencephalic trigeminal nucleus of adult rat in vitro.

The mesencephalic trigeminal nucleus (MesV) contains the somata of primary afferent neurons innervating masticatory muscle spindles and the periodontal membrane. MesV afferent somata are unique in receiving synaptic inputs. Intracellular recordings in coronal pontine slices from adult rats were made from MesV neurons identified by having Cs-sensitive inward rectification and pseudounipolar morphology. Stimuli near the MesV evoked either a cluster of action potentials superimposed on a postsynaptic potential (PSP) or an antidromic spike at resting membrane potential (RMP). Membrane hyperpolarization revealed that each cluster of action potentials consisted of an antidromic spike and a subsequent PSP. Evoked PSPs in slices and miniature postsynaptic currents (mPSCs) recorded using whole-cell patch in dissociated MesV neurons were resistant to glutamate antagonists and strychnine but were reversibly abolished by 40 microM bicuculline. Superfusion of 1-10 mM GABA decreased input resistance and depolarized the membrane. Reversal potentials for evoked PSPs and GABA-induced depolarizations were similar and close to that for mPSCs which matched the Cl- equilibrium potential. Thus activation of synapses on MesV somata evokes GABAergic PSPs that generate action potentials at RMP in the adult. These data also indicate that primary afferent MesV neurons can act as interneurons in the central control of mastication.

Anti-angiogenic potential of tocotrienol in vitro.

Modulation of angiogenesis is now a recognized strategy for the prevention of various angiogenesis-mediated disorders. We investigated, using well-characterized in vitro systems, the anti-angiogenic property of vitamin E compounds, with particular emphasis on tocotrienol, a natural analog of tocopherol. Tocotrienol, but not tocopherol, inhibited the proliferation of bovine aortic endothelial cells in dose dependent manner at half-maximal concentrations in the low micromolar range. Tocotrienol also significantly inhibited the formation of networks of elongated endothelial cells within 3D collagen gels. From these results, we suggest that tocotrienol is a potential candidate for the development of useful therapeutic agents or preventive food factors for tumor angiogenesis.

Effect of different proton pump inhibitors, differences in CYP2C19 genotype and antibiotic resistance on the eradication rate of Helicobacter pylori infection by a 1-week regimen of proton pump inhibitor, amoxicillin and clarithromycin.

AIM: To investigate the effect of different proton pump inhibitors, S-mephenytoin 4'-hydroxylase (CYP2C19) genotype and antibiotic susceptibility on the eradication rate of Helicobacter pylori. METHODS: One hundred and eighty-seven H. pylori-infected peptic ulcer patients were randomly treated with either rabeprazole (10 mg b.d.) or lansoprazole (30 mg b.d.) plus amoxicillin (750 mg b.d.) and clarithromycin (400 mg b.d.) for 1 week. The antibiotic susceptibility and CYP2C19 genotype (extensive or poor metabolizer) were investigated. RESULTS: The eradication rates in the rabeprazole-amoxicillin-clarithromycin (RAC) and lansoprazole-amoxicillin-clarithromycin (LAC) groups were 75% and 69%, respectively, on an intention-to-treat basis, and 80% and 75%, respectively, on a per protocol basis. The eradication rate for clarithromycin-resistant strains was significantly lower than that for clarithromycin-sensitive strains (24% vs. 86%, P < 0.05). For clarithromycin-sensitive strains in the LAC group, there was a tendency for a lower eradication rate in extensive than poor metabolizers. The eradication rate in extensive metabolizers in the RAC group tended to be higher than that in extensive metabolizers in the LAC group (89% vs. 78%, P = 0.079726). CONCLUSIONS: The success of the 1-week proton pump inhibitor-amoxicillin-clarithromycin regimen depends on the susceptibility of H. pylori to clarithromycin. Moreover, differences in CYP2C19 genotype influence the eradication rates of lansoprazole-based therapy, and the rabeprazole-based regimen has an advantage especially in extensive metabolizers.

Cloning and expression of a Porphyromonas gingivalis gene for protoporphyrinogen oxidase by complementation of a hemG mutant of Escherichia coli.

Porphyromonas gingivalis, a bacterium implicated in periodontal pathogenesis, has a growth requirement for iron protoporphyrin IX. By complementation with a P. gingivalis 381 chromosomal DNA library, we were able to isolate a clone that enhanced the poor growth of a hemG mutant of Escherichia coli. The DNA sequence analysis of this clone revealed three open reading frames (ORFs). ORF3 encoded a protein of 466 amino acids with a calculated molecular weight of 51 695 Da. The deduced amino acid sequence of the ORF3 gene had significant similarity to sequences of protoporphyrinogen oxidase (PPO) from Myxococcus xanthus (30% identical residues). When the ORF3 gene was overexpressed in E. coli, the extract had much higher PPO activity than a control extract, and this activity was inhibited by acifluorfen, a specific inhibitor of PPO. Thus, ORF3 was named PgHemG. Furthermore, several porphyrin-related genes, including hemD, hemN and hemH, were identified in the data bases on the websites available on-line. We postulated that a porphyrin biosynthetic pathway to heme from preuroporphyrin may be conserved in P. gingivalis.

Proliferation and functional changes of pancreatic gastrin cells in neonatal rat.

INTRODUCTION: Although gastrin cells are not found in the adult pancreas, they are found transiently in the neonatal pancreas. It has been suggested that gastrin may play a role in pancreatic development. However, cell kinetics as well as the fate and the role of gastrin cells are not clear. METHODOLOGY: Proliferation and functional changes of pancreatic gastrin cells in neonatal Wister rats were studied by immunohistochemistry and [(3)H]thymidine autoradiography. RESULTS: Numbers of pancreatic gastrin cells in neonatal rats showed a peak immediately after birth and then decreased rapidly. Gastrin cells were observed within approximately 2 weeks after birth in islets and within approximately 4 weeks after birth among exocrine cells. In contrast with the decrease of gastrin cell numbers, numbers of duodenal cholecystokinin cells increased remarkably after 7 days of age. Proliferative activity of acinar cells showed two peaks at age 2 days and 9 days. Despite a decrease in gastrin cell numbers, gastrin cells maintained a certain degree of proliferative activity. The "re-staining method" for gastrin and insulin revealed that immunoreactive cells for both gastrin and insulin were rarely found a few days after birth. CONCLUSION: These results suggest that pancreatic gastrin cells do not die off or change to another type of endocrine cell and that some gastrin cells change to insulin cells.

Correlation between electrophysiology and morphology of three groups of neuron in the dorsal commissural nucleus of lumbosacral spinal cord of mature rats studied in vitro.

The dorsal commissural nucleus (DCN) in the lumbosacral spinal cord receives afferent inputs from the pelvic organs via pudendal and pelvic nerves. Electrophysiological and morphological properties of neurons in the DCN of L6-S1 were examined using whole-cell recordings with biocytin-filled electrodes in transverse slices of mature rat spinal cord. Neurons were categorized into three groups according to their discharge in response to suprathreshold depolarizing pulses; neurons with tonic (19/42) and phasic (13/42) firing patterns, and neurons (10/42) that fired in bursts arising from a Ca(2+)-dependent hump. The predominantly fusiform somata of neurons labeled during recording (n = 31) had on average 3.1 primary dendrites, 7.5 terminating dendritic branches, 3.1 axon collaterals, and 14.2 axon terminations per neuron. The groups were morphologically distinct on the basis of their dendritic branching patterns. Phasic neurons (n = 10) had the most elaborate dendritic branching and the largest numbers of axon collaterals. All tonic neurons (n = 11) had axons/collaterals projecting to the intermediolateral area but none to the funiculi, suggesting that they function as interneurons in local autonomic reflexes. Many axons/collaterals of all phasic neurons lay within the DCN, suggesting that they integrate segmental and descending inputs. Seven of 10 neurons with Ca(2+)-dependent humps had axons/collaterals extending into one of the funiculi, suggesting that they project intersegmentally or to the brain. Ca(2+) hump neurons also had more axons/collaterals within the DCN and fewer in the intermediolateral area than tonic neurons. This correlation between firing pattern and morphology is an important step toward defining the cellular pathways regulating pelvic function.

Temperature-sensitive mutations in various genes of Escherichia coli K12 can be suppressed by the ssrA gene for 10Sa RNA (tmRNA).

An Escherichia coli strain with a deletion in the ssrA gene that encodes 10Sa RNA (tmRNA) was used to screen for temperature-sensitive (ts) mutants whose ts phenotypes were suppressible by introduction of the wild-type ssrA gene. Mutants in four different genes were isolated. Ts mutants of this type were also obtained in a screen for mutations in thyA, the structural gene for thymidylate synthase. The ThyA activity in crude extracts prepared from the ts mutants was temperature-sensitive. The presence of the ssrA gene caused an increase in the total amount of the temperature-sensitive enzyme expressed, rather than suppressing the ts activity of the enzyme itself. SsrA-DD, a mutant form of 10Sa RNA, suppressed the ts phenotype of a thyA mutant, suggesting that degradation of a tagged peptide was not required for suppression of the ts phenotype. Considering the fact that ssrA-suppressible mutants could be isolated as temperature-sensitive mutants with mutations in different genes, it seems evident that trans-translation can occur on mRNA that is not lacking its stop codon.

Ca2+-independent phospholipase A2 activity in apical plasma membranes from the rat parotid gland.

An apical-enriched plasma membrane fraction (A-PM) was prepared from rat parotid gland by Mn2+ precipitation. In this fraction, phosphatidylcholine (PC) labelled at the sn-2 position was mainly decomposed into two labelled compounds (free fatty acid and 1,2-diacylglycerol) under Ca2+-free conditions. Studies using double-labelled PC and 2,3-diphosphoglycerate (as a phospholipase D inhibitor) showed that they were produced through different pathways: free fatty acid was released by phospholipase A2 (PLA2) while 1,2-diacylglycerol may be produced by sequential action of phospholipase D and phosphatidate phosphatase. The PLA2 in A-PM did not require Ca2+ for its activity and was highly activated by Triton X-100 and ATP. The inhibitor of the well-documented Ca2+-independent PLA2, bromoenol lactone, did not inhibit the PLA2 activity in A-PM. Although PLA2 activity was detected in other subcellular fractions, the highest specific activity was in A-PM. Its distribution among various fractions was roughly similar to that of the marker enzyme of apical plasma membranes. These findings suggested that Ca2+-independent PLA2 activity is present in apical plasma membranes from rat parotid gland. In addition, to clarify the involvement of the PLA2 in exocytosis, the fusion of exogenous PLA2-treated membranes with secretory granules was examined by fluorescence dequenching assay. This study clearly demonstrated the facilitation of fusion by PLA2 treatment, which suggests some involvement of apical PLA2 in saliva secretion.