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Biomedical subjects

H C Chung

Publications and source records attributed to H C Chung.

At least 19 recordsLinked to original sources
PubMed1998

Synchronous elevation of soluble intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) correlates with gastric cancer progression.

Soluble forms of ICAM-1 (sICAM-1) and VCAM-1 (sVCAM-1) have been reported from the supernatant of cytokine-activated endothelial cells, cancer cells and from sera of cancer patients. We measured sICAM-1 and sVCAM-1 from the serum of 20 healthy volunteers and 142 gastric cancer patients by ELISA assay. Ninety-five patients were operable and 47 patients were in-operable at the time of this study. Particularly in the 28 operable patients, we sampled both portal and peripheral blood simultaneously and measured the levels of the soluble forms of cell adhesion molecules (sCAMs). The sCAMs level and sero-positivity rate increased with cancer progression in order of the healthy controls, operable patients, and inoperable patients. In in-operable cancer, the sICAM-1 level increased more with liver metastasis. sICAM-1 and sVCAM-1 did not correlate with each other in either portal or peripheral blood. A total of 58.3% of patients with liver metastasis and 22.9% of patients without liver metastasis showed synchronous expression of both sCAMs (p = 0.03). Synchronous sero-positivity of sCAMs and alpha FP was higher with liver metastasis (p = 0.01). The median overall survival duration which co-expressed both sCAMs was 9 months. This showed a significant difference compared with the sICAMs non-expressing group, where the median survival was not reached until 24 months follow-up (p = 0.002). The synchronous expression of sCAMs was an independent risk factor in gastric cancer patients. We raise the possibility that synchronous sICAM-1 and sVCAM-1 elevation may be a useful monitor to determine tumor burden in gastric cancer.

Adult↗
PubMed1997

Comparison of biological phenotypes according to midkine expression in gastric cancer cells and their autocrine activities could be modulated by pentosan polysulfate.

We studied biological phenotypes of gastric cancer cell lines based on a novel heparin-binding growth/differentiation factor (midkine (MK)) expression. MK expression was found in 67% (6/9) of the gastric cancer cell lines and 56% (14/25) of the primary cancer tissues. Gastric cancer cell lines with MK expression showed higher colony forming activity in soft agar assay and endothelial cell growth stimulatory effect in cross-feeding assay than cells which did not express MK. However, urokinase-type plasminogen activator (uPA) expression and tumor invasiveness did not correlate with MK expression. Growth of MK expressing cells was inhibited by a heparin-binding blocking agent, pentosan polysulfate (PPS). In cancer tissues, MK expression correlated with tumor size, suggesting in vivo autocrine and paracrine activity. This proliferation promoting activity of MK can be targeted by an anti-heparin binding agent as a biotherapy model in gastric cancer.

Ascitic Fluid↗
PubMed1997

High level of urokinase-type plasminogen activator is a new prognostic marker in patients with gastric carcinoma.

BACKGROUND: Prognosis of gastric carcinoma is related to invasion and metastasis. Evidence has accumulated that invasion and metastasis in solid tumors require the action of tumor-associated proteases, which promote the dissolution of the surrounding tumor matrix and the basement membrane. The serine protease urokinase-type plasminogen activator (uPA) and its inhibitor, plasminogen activator inhibitor-1 (PAI-1), appear to have a major function in these processes. Recent reports have demonstrated that expression of these proteolytic enzymes is elevated in breast and colon carcinoma and that it can be associated with invasiveness and poor prognosis. Therefore, the authors evaluated whether the expression and activation of uPA and PAI-1 might be of clinical value as a tumor/biologically defined risk factor in patients with gastric carcinoma. METHODS: Enzyme-linked immunoadsorbent assays were used to test for uPA antigens and PAI-1 in tissue extracts of normal and cancerous tissue from 160 gastric carcinoma patients who were enrolled in the Yonsei Cancer Center Study Group. RESULTS: Both uPA and PAI-1 levels were significantly higher in cancerous tissues than in normal tissues (uPA: 9.4 +/- 8.7 vs. 5.3 +/- 3.1 ng/mg protein cytosol; PAI-1: 10.9 +/- 9.1 vs. 5.8 +/- 2.9 ng/mg protein cytosol), (P < 0.001, respectively). Both high uPA and PAI-1 levels were associated with differentiation of the tumor (P = 0.04 and P = 0.004, respectively), and a high PAI-1 level was associated with lymph node metastasis at an advanced stage (P = 0.003 and P = 0.04, respectively). There was a correlation between the levels of uPA and PAI-1 expression in cancerous tissues (correlation coefficient = 0.57). In univariate analysis, a high level of uPA or PAI-1 was associated with a short relapse free survival, but in multivariate analysis only a high level of uPA was an independent prognostic parameter for a short relapse free survival for gastric carcinoma patients. CONCLUSIONS: These data indicate that uPA is a new independent variable for the identification of high risk gastric carcinoma patients. Therefore, therapy targeting uPA can be applied as a new biologic treatment modality for these individuals.

Adult↗
PubMed1997

Correlation between K-ras gene mutation and prognosis of patients with nonsmall cell lung carcinoma.

BACKGROUND: Mutations at codons 12, 13, and 61 of the three ras genes, H-ras, K-ras, and N-ras, convert these genes into active oncogenes. It appears that ras gene mutations can be found in a variety of tumor types. The purpose of this study was to evaluate the clinical significance of K-ras gene mutation in nonsmall cell lung carcinoma (NSCLC). METHODS: The authors analyzed 58 NSCLC patients for mutations at codons 12, 13, and 61 of the K-ras gene and correlated the findings with the tumor stage and patient survival. Polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) and the direct nucleotide sequencing method were used to detect mutations after amplification of ras specific sequences by PCR. RESULTS: Fourteen mutations (24%) of ras genes were found, all at codon 12 of the K-ras gene. GGT to GAT transition was the predominant mutational pattern. There was a significant association between K-ras mutation and the tumor stage (i.e., the higher the stage, the higher the mutation rate) (P = 0.014). Using univariate analysis, the presence of K-ras mutation in paraffin embedded tissue from patients who received treatment with curative intent was associated with a shorter survival (P = 0.039). The median survival duration for patients with or without K-ras mutation was 9 and 30 months, respectively. The Cox proportional hazards model also predicted a higher risk for patients with K-ras mutations (P = 0.047). CONCLUSIONS: K-ras mutations, present in a subset of NSCLC, are associated with tumor progression and shortened patient survival.

Adult↗
PubMed1997

Sequential production and activation of matrix-metalloproteinase-9 (MMP-9) with breast cancer progression.

The degradation of the basement membrane by matrix-metalloproteinase (MMP) and serine protease is a critical point in tumor invasion and metastasis. We measured the activity of MMP-9 from 28 normal, 12 benign and 126 breast cancer tissues using gelatin zymography with an image analysis system. ProMMP-9 was expressed in 17.5% of the cancer patients compared to 2.5% in 40 non-cancerous tissues (p = 0.014). The mature form of MMP-9 (82 kD) was expressed only in T2-T4 stages. During the early phase of breast cancer (DCIS and T1 stage) progression, only production of proMMP-9 increased. However, as the cancer grew or invaded skin (T2-T4), or with lymphovascular permeation, both production and activation of MMP-9 increased. In conclusion, proMMP-9 production was the main cause of increased MMP-9 activity during the early phase, while both production and activation increased in the late phase of breast cancer.

Breast Neoplasms↗
PubMed1997

P-glycoprotein: the intermediate end point of drug response to induction chemotherapy in locally advanced breast cancer.

Expression and clinical relevance of p-glycoprotein (p-gp) were evaluated in 31 cases of locally advanced breast cancer and 9 cases involving inflammatory breast cancer after induction chemotherapy. The de novo p-gp expression rate was 26% and increased up to 58% (p = 0.03) with the FAC (5-fluorouracil, adriamycin, cyclophosphamide) regimen. Although more clinically complete responders were found in the secondary p-gp negative group (p = 0.02), this difference was not found in pathological tumor response. Moreover, as the grade of the secondary p-gp expression increased, the chemotherapeutic effect decreased, suggesting an inverse relationship between p-gp expression and drug effect (p = 0.04). When we subgrouped the patients into 4 groups using these two parameters, p-gp negative patients presenting with a high drug effect showed a low recurrence rate (p = 0.05) and marginal survival benefits (p = 0.09) as opposed to patients with a low drug effect. But in p-gp positive groups, the recurrence rate was the same between the two groups regardless of the drug effect. Thus, in the p-gp negative patient with a high drug effect, adjuvant chemotherapy with the same regimen as induction chemotherapy may induce more prognostically favorable results. Therefore, clinical application of the secondary p-gp detection can be used as an intermediate endpoint in evaluating drug response for an induction regimen.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1997

Retrospective comparison of infusional 5-fluorouracil, doxorubicin, and mitomycin-C (modified FAM) combination chemotherapy versus palliative therapy in treatment of advanced gastric cancer.

About one-third of patients with gastric cancer are unresectable at the time of diagnosis. Their median survival is < 6 months, with a grave prognosis. The purpose of this study was to assess the efficacy of a modified FAM (mFAM) regimen in advanced gastric cancer. We retrospectively reviewed the clinical records of 409 advanced gastric cancer patients who had not received curative surgery. Among 409 patients, 202 patients were treated with an mFAM regimen (infusional 5-FU + doxorubocin + mitomycin-C), and 207 patients received no chemotherapy (control group). No differences were found in clinical parameters between the two groups. The 1-year survival rates were 34.1% for the mFAM-treated group and 22.5% for the control group (p = 0.0135). In subset analysis, a higher 1-year survival rate was demonstrated in patients with mFAM and palliative surgery. Of the 154 evaluable patients in the mFAM-treated group, the response rate was 17.5%. In these patients, median response duration was 30 weeks, and progression-free survival was 23 weeks. Overall toxicity of mFAM regimen was relatively tolerable and reversible. In conclusion, FAM combination chemotherapy, which has been used as a standard therapy, prolonged survival after modification of the administration schedule and combination with palliative surgery. A prospective randomized study is warranted to confirm this conclusion from our retrospective study.

Adenocarcinoma↗
PubMed1997

A comparative study of intravenous granisetron versus intravenous and oral ondansetron in the prevention of nausea and vomiting associated with moderately emetogenic chemotherapy.

We conducted a prospective, randomized, open, single-center, parallel group study comparing the anti-emetic efficacy and toxicity of granisetron with that of ondansetron in patients receiving moderately emetogenic chemotherapy. From December 1994 to May 1995, patients who were to receive moderately emetogenic chemotherapy for the first time or who had not received chemotherapy (80 to 100 mg/m2 of cisplatin or 40 mg/m2 of doxorubicin) within 4 weeks previously were enrolled in this study. The following anti-emetic regimens were used: 3 mg of granisetron were given intravenously before chemotherapy for a single dose; 8 mg of ondansetron were given intravenously before chemotherapy and then every 8 hours for a total of 3 doses, plus 8 mg of an oral maintenance dose every 12 hours for 5 consecutive days. We evaluated 97 patients (48 received granisetron and 49 received ondansetron). In the first 24 hours after chemotherapy, complete and major responses were achieved in 76.6% of the patients receiving granisetron and in 72.9% of patients receiving ondansetron (p = 0.9033). Additionally, there was no difference in the control of delayed nausea and vomiting between the two groups (51.1% versus 54.2%, p = 0.9200), and there were no significant adverse effects or toxicities. We have concluded that a single dose of granisetron is as effective in prophylaxis of emesis induced by moderately emetogenic chemotherapy as a triple dose of ondansetron plus oral maintenance.

Administration, Oral↗
PubMed1997

Comparison of epidural butorphanol plus clonidine with butorphanol alone for postoperative pain relief.

BACKGROUND: Epidural butorphanol has been shown to produce effective analgesia with less side effects than that of morphine but relatively short duration. Clonidine, an alpha 2-adrenergic agonist, has been reported to provide [corrected] pain relief by epidural administration. Furthermore, epidural clonidine has been shown to potentiate the analgesic effect of epidural morphine. The present study was undertaken to evaluate the analgesic and side effects of epidural administration (Ep) of butorphanol and clonidine. METHODS: After giving their consents, 60 adult patients scheduled for abdominal surgeries were enrolled in this study. Prior to anesthesia induction, indwelling lumbar epidural catheters were placed in all patients who then received general anesthesia with inhalation anesthetic without narcotic analgesics. In the postoperative period, when the patients first complained of pain, they were divided into 2 equal groups of 30 patients each in a randomized and double blinded fashion with Group I receiving Ep butorphanol 0.5 mg and Group II receiving Ep butorphanol 0.5 mg plus clonidine 75 micrograms. All patients were observed for pain relief, sedation, vital signs, arterial blood gas studies and adverse effects for 12 h. RESULTS: Onset of pain relief with epidural butorphanol began at 5 min and peaked at 20-30 min with a duration of action lasting 4-6 h. The combination of butorphanol and clonidine had numerically superior pain relief than that of butorphanol for the first 30 min but it did not attain statistical significant difference. The duration of action with the combination group was similar to that of butorphanol alone. Incidence of adverse effects were similar in both groups except that hypotension and more pronounced sedation were observed in Group II. CONCLUSIONS: Our study showed that the addition of clonidine to epidural butorphanol did not enhance its analgesic effect in any significant manner nor did it reduce the adverse effects. This combination does not seem to offer any advantage for clinical use.

Adrenergic alpha-Agonists↗
PubMed1997

Intermittent bolus versus patient-controlled epidural morphine for postoperative analgesia.

BACKGROUND: Patient-controlled epidural analgesia (PCEA) is a technique that combines the flexibility and convenience of PCA with the intrinsic analgesic efficacy of epidurally administered opioids. The aim of this study is to compare the analgesic and side effects of intermittent bolus injections of epidural morphine with PCEA using morphine during the first 24 h after elective low abdominal surgery. METHODS: Sixty patients were randomly assigned into two groups. Patients in group I (n = 30) received 3 mg epidural morphine at an 8-hour interval after surgery. Patients in group II (n = 30) received 2 mg epidural morphine after surgery and patient-controlled analgesia device was processed to deliver morphine 0.15 mg/h, 0.15 mg/bolus. All patients were observed for pain relief and adverse effects for 24 h. RESULTS: Mean morphine consumption was 9 mg for epidural morphine group and 6.87 +/- 0.27 mg for PCEA group. Although the PCEA group utilized significantly less morphine (p < 0.05), pain and sedation scores were similar in the two groups. Pruritus occurred more frequently in the epidural morphine group (63%) than in the PCEA group (33%). Frequency and severity of nausea and vomiting were similar in the two groups. CONCLUSIONS: PCEA with morphine decreases morphine consumption and with less adverse effects than intermittent bolus of epidural morphine. PCEA with morphine is an acceptable alternative to epidural morphine after low abdominal surgery.

Analgesia, Epidural↗
PubMed1997

Retention of broken central venous catheters in pulmonary artery and inferior vena cava--a case report.

Central venous catheterization (CVC) has become an important maneuver both for measuring the central venous pressure and for carrying out long-term intravenous alimentation. Furthermore, a central vein may be needed for the rapid restoration of blood volume in condition of acute hemorrhage with difficulties establishing of a peripheral cannulation. However, complications related to either the venipuncture or the presence of catheters in the central venous system were reported. The case we reported here had two ruptured catheter fragments retained in the great vessels, one in the right subclavian vein and the other one in the femoral vein during CVC. They migrated to the pulmonary artery and inferior vena cava, and were finally removed through open sternotomy with pulmonary arteriotomy and exploration of common iliac vein two weeks later. This was an unusual and possibly fatal complication.

Adult↗
PubMed1996

Sequential activation and production of matrix metalloproteinase-2 during breast cancer progression.

The proteolytic processes are thought to be the critical point in tumor invasion and metastasis, mainly by matrix metalloproteinases (MMPs) and serine proteases. We measured the activity of MMP-2 from 28 normal, 12 benign and 126 breast cancer tissues using gelatin zymography. Inactive MMP-2 (72 kD) was expressed in 53.6% of the normal and 66.6% of the cancer tissues, respectively (P = 0.77), while active MMP-2 (62 kD) was expressed in 28.6% and 73.0%, respectively (P = 0.003). The enzymatic activity of active MMP-2 (62 kD) measured in the gel band area was 4.0 +/- 7.2 mm2 in normal breasts, 7.7 +/- 9.8 mm2 in benign breast diseases, 9.5 +/- 8.5 mm2 in ductal carcinoma in situ (DCIS), and 12.0 +/- 13.7 mm2 in invasive cancers. The MMP-2 activation ratio (62 kD/62 kD + 72 kD) was 0.12 +/- 0.18 in normal tissues, 0.10 +/- 0.20 in benign diseases, 0.61 +/- 0.22 in DCIS, and 0.50 +/- 0.28 in invasive cancers. In conclusion, MMP-2 activation was the main cause of the increased 62 kD MMP-2 activity during the early phase of breast cancer, while production of MMP-2 supplemented the increased 62 kD activity in the late phase. We suggest, therefore, that these differential expressions of MMP-2 activation and production during the different stages of breast cancer progression are potential therapeutic targets for biological or gene therapy under the concept of stage-oriented cancer treatment.

Breast↗
PubMed1996

Gastric cancer in young patients who underwent curative resection. Comparative study with older patients.

Although several studies demonstrated the poor prognosis of young patients with gastric cancer, there were few reports about the survival of young patients who underwent radical surgery. We retrospectively reviewed the hospital records of 697 patients with curatively resected locally advanced gastric cancer to compare the pathologic findings and prognosis of young patients (aged < 40 years; 91 patients) with those of older (aged > or = 40 years; 606 patients) patients. There were no significant differences in location of tumors, Borrmann types, tumor invasion, and pathologic stage between both age groups. Despite the male predominance in older patients, the male-to-female ratio was nearly equal in young patients (p = 0.022). A significantly higher percentage of young patients showed poorly differentiated histology including signet ring cell and undifferentiated carcinoma than older patients (p < 0.0001). Adjuvant chemotherapy was more frequently performed in the young patients with a variety of different regimens (85.7 vs. 72.3%; p = 0.009). The 5-year overall survival rate revealed no statistically significant differences between the young and older patients (53.1 vs. 56.6%; p = 0.820). In conclusion, the young patients with curatively resected locally advanced gastric cancer showed similar survival compared to that of older patients.

Adult↗
PubMed1996

P-glycoprotein as an intermediate end point of drug resistance to neoadjuvant chemotherapy in locally advanced gastric cancer.

The expression of p-glycoprotein (p-gp) was evaluated in pre- and post-chemotherapy states after the administration of adriamycin-based chemotherapy in 24 gastric cancer patients. Among them, group A was composed of twelve patients who relapsed after surgery plus adjuvant chemotherapy and group B was composed of another twelve patients who received neoadjuvant chemotherapy plus surgery. Pre-chemotherapy p-gp was evaluated in 18 out of 24 patients (6 patients had no pre-chemotherapy paraffin blocks) and post-chemotherapy p-gp was evaluated from all 24 patients. Pre- and post-chemotherapy p-gp was expressed in 5 of 18 patients (27.8%), and 9 of 24 patients (37.5%), respectively, with immunohistochemical stain using monoclonal antibody JSB-1. No differences of disease-free survivals were observed in Group A based on post-chemotherapy p-gp expression from relapsed lesions. In Group B, there was a higher relapse rate (p = 0.04) and a lower one-year disease-free survival rate (p = 0.04) in post-chemotherapy p-gp positive patients when adjuvant treatment was done with the same regimen as neoadjuvant chemotherapy. In all patients studied, post-chemotherapy p-gp expression correlated with a higher systemic recurrence (p = 0.04). These data suggest that p-gp can be induced by an adriamycin-based chemotherapy in gastric cancer. Thus, we suggest that the prognosis of gastric cancer may be poor if a multidrug resistance (MDR)-related regimen is used in the presence of p-gp after neoadjuvant chemotherapy with an adriamycin-based regimen, even if the initial response is good.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1996

Molecular mechanism for alkyl sulfide-modulated carbon tetrachloride-induced hepatotoxicity: the role of cytochrome P450 2E1, P450 2B and glutathione S-transferase expression.

The modulation of CCl4-induced hepatotoxicity in response to alkyl sulfides and alkyl ethers including allyl disulfide (ADS), allyl sulfide (AS), allyl ether (AE), propyl disulfide (PDS), propyl sulfide (PS), propyl ether (PE) and butyl sulfide (BS) was studied. Whereas pretreatment of rats with either ADS or AS (50 mg/kg, 7 days) blocked a CCl4-induced increase in plasma alanine aminotransferase (ALT) activity by 91 and 56%, respectively, AE, PDS, PS, PE or BS treatment enhanced CCl4-induced ALT activity by 52, 55, 238, 25 or 86%, respectively. Histochemical examinations supported the results of plasma ALT activity. Injection of GdCl3 to PS-pretreated rats failed to block the potentiated ALT increase, whereas GdCl3 completely prevented vitamin A-enhanced elevation of ALT activity. AS treatment completely blocked PS-potentiated CCl4 intoxication. Concomitant treatment of animals with both PS and vitamin A followed by a CCl4 insult resulted in super-potentiation of CCl4-induced hepatotoxicity, suggesting that the mechanism of PS-enhanced hepatotoxicity differs from that caused by vitamin A. Pyridine or phenobarbital potentiation of CCl4-induced increases in ALT activity implys that cytochrome P450 2E1 (P450 2E1) and P450 2B expression may be associated with the increased toxicity. P450 2E1 expression appeared to be associated with the alkyl sulfide-modulated hepatotoxicity, as evidenced by both immunoblot analyses and metabolic activity. P450 2B immunoblot analysis revealed that either AS or PS substantially induced hepatic P450 2B1/2 levels. Thus, PS-enhanced CCL4 hepatotoxicity may be related in part with P450 2B induction. ADS, AS or PS treatment caused increases in the glutathione S-transferase (GST) conjugating activity toward 1-chloro-2,4-dinitro-benzene. ADS, AS or PS induced Ya and Yb1 subunits by 2- to 3-fold. ADS or AS treatment also significantly elevated the levels of Yc subunits. PS failed to induce Yc expression, although this agent effectively increased Yb2 expression. Northern blot analyses revealed that ADS and AS concomitantly stimulated GST Ya, Yb1 and Yc2 gene expression, whereas PS increased the levels of Ya, Yb1, and Yb2 mRNA, but not Yc2 mRNA levels. The expression of GST subunit Yc2 in response to these compounds might be associated with hepatoprotective effects. These results demonstrate that ADS and AS have distinct capability of blocking CCl4-induced hepatotoxicity, whereas certain saturated alkyl sulfides rather potentiate CCl4-induced hepatotoxicity and that the underlying mechanism is associated with P450 2E1 and P450 2B expression, and possibly with certain GST expression.

Allyl Compounds↗
PubMed1995

Regulation of phosphatidylinositol kinases by arachidonic acid in rat submandibular gland cells.

Phosphoinositide kinases were characterized in membrane extracts of rat submandibular gland cells. Both phosphatidylinositol (PI) 4-kinase and phosphatidylinositol-4-phosphate (PI(4)P) 5-kinase phosphorylated endogenous substrates in reactions that were linear for up to 5 min, were activated by Mg2+ and showed maximal activity around neutral pH. PI 4-kinase was stimulated by Triton X-100 at an optimal concentration of 0.22%, but the detergent had an inhibitory effect on PI(4)P 5-kinase. Arachidonic acid (AA), at concentrations greater than 100 microM, inhibited the activity of both enzymes in a dose-dependent manner. The inhibitory effect was replicated by other unsaturated fatty acids, but not by a saturated fatty acid of the sn-20 series. The nature of AA inhibition of the kinases was examined in enzyme kinetic studies with exogenous phosphoinositide and adenosine 5'-triphosphate (ATP) substrates. Lineweaver-Burk plots of PI 4-kinase activity showed that AA had no effect on the apparent Km for either PI or ATP, but that the fatty acid significantly reduced Vmax (PI) from 331 to 177 pmol.mg-1.min-1 and Vmax (ATP) from 173 to 59 pmol.mg-1.min-1. This inhibitory action was consistent for PI(4)P 5-kinase kinetics, where again, AA did not alter apparent Km values, but lowered Vmax for both PI(4)P and ATP by around 50%. Since the combination of a reduced Vmax and an unchanged Km value indicates noncompetitive enzyme inhibition, it is proposed that AA regulates phosphoinositide cycle activity in submandibular gland cells by acting as a noncompetitive inhibitor of PI 4-kinase and PI(4)P 5-kinase.

1-Phosphatidylinositol 4-Kinase↗
PubMed1995

Muscarinic regulation of phospholipase D and its role in arachidonic acid release in rat submandibular acinar cells.

The characteristics of muscarinic cholinergic-induced phospholipase D (PLD) activation, and the involvement of the enzyme in the release of arachidonic acid were examined in rat submandibular acinar cells. Carbachol produced a dose-related activation of PLD to around fivefold control values at 100 microM agonist concentration. This was associated with the appearance of free choline, phosphatidic acid and arachidonic acid, indicating that the PLD substrate was phosphatidylcholine. The response to carbachol was inhibited by 60% by U73122, a blocker of a phospholipase C (PLC) specific to phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P2], suggesting that the cleavage of phosphatidylcholine by PLD was, at least in part, secondary to agonist-coupled hydrolysis of PtdIns(4,5)P2 by PLC. Consistent with this, PLD was also activated to levels comparable to those induced by carbachol, by the phorbol ester, 12-O-tetradecanoylphorbol-13-acetate (TPA), and the Ca2+ mobilizer, thapsigargin, two agents that respectively mimic the activation of protein kinase C (PKC) by diacylglycerol and the elevation of cytosolic Ca2+ by inositol 1,4,5-triphosphate [Ins(1,4,5)P3] in the phosphoinositide effect. The cell-permeant Ca2+ chelator 1,2-bis-(O-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid, tetraacetoxymethyl ester (BAPTA/AM) abolished the thapsigargin-induced activation of PLD and inhibited the responses of PLD to carbachol and TPA by 60%. The PKC inhibitor, Ro-31-8220, also inhibited the activation of PLD by carbacol and TPA to a level of approximately double control values, but had no effect on the thapsigargin-induced elevation of PLD. A role for both the PKC-associated and Ca(2+)-mobilizing arms of the PtdIns(4,5)P2-PLC pathway in PLD regulation is thus suggested. Pretreatment of cells with the phosphatidate phosphohydrolase blocker, propranolol, significantly enhanced the carbachol-induced elevation of phosphatidic acid, but decreased agonist-stimulated production of diacylglycerol and arachidonic acid, indicating that phosphatidlycholine was the likely source of arachidonic acid. We therefore propose that, in submandibular mucous acinar cells, muscarinic activation of the PtdIns(4,5)P2-PLC pathway regulates phosphatidylcholine-specific PLD through both the PKC- and Ca(2+)-mobilizing arms of the phosphoinositide response, and that diacylglycerol, derived from phosphatidylcholine via phosphatidic acid, is a source of free arachidonic acid.

Animals↗