Biomedical subjects

R J Bold

Publications and source records attributed to R J Bold.

At least 19 recordsLinked to original sources

PubMed1998

A human gastric cancer cell line possesses a functional receptor for gastrin-releasing peptide.

Bombesin (BBS) exhibits diverse biological functions including those of neurotransmitter, regulator of gastrointestinal hormone release, and mitogen. Gastrin-releasing peptide (GRP, the mammalian equivalent of BBS) is found in mucosal cells of the gastric fundus and antrum. We determined whether a human gastric cancer cell line (SIIA) expresses a functional GRP-receptor (GRP-R). BBS increased intracellular calcium ([Ca2+]i), and a specific GRP-R antagonist, ([D-Phe6, Des-Met14]-BBS (6-14)-ethylamide), blocked BBS-induced increase in [Ca2+]i. SIIA cells possess GRP-R mRNA by reverse transcriptase-PCR. Furthermore, these cells possess an 80-kDa cell surface protein that specifically binds BBS with two high-binding affinities (Kd1 = 0.6 nM, Kd2 = 6.7 nM). These findings indicate that SIIA cells possess a GRP-R that is capable of physiological signal transduction, though the cellular response remains unknown.

Adenocarcinoma↗

PubMed1996

Surgery.

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Bacterial Translocation↗

PubMed1996

Hyperphosphorylation of retinoblastoma protein and stimulation of growth by okadaic acid in human pancreatic cancer.

Phosphorylation/dephosphorylation of intracellular proteins are important steps in the regulation of cell growth. Okadaic acid, an inhibitor of the serine/threonine protein phosphatases 1 and 2A, is a potent tumor promoter. This effect may be through the inhibition of dephosphorylation (termed "hyperphosphorylation") and subsequent inactivation of tumor-suppressor proteins. We examined whether okadaic acid regulates growth of human pancreatic cancer cells (MIA PaCa-2 and Panc-1) or alters the phosphorylation of the retinoblastoma tumor-suppressor protein. Growth studies, nuclear labeling analyses, and Western blotting for retinoblastoma protein were performed. Okadaic acid stimulated cell growth and induced hyperphosphorylation of the retinoblastoma protein. The growth-stimulatory effect of okadaic acid on these human pancreatic cancer cells may be mediated by inactivation of the growth suppressive effect of the retinoblastoma protein by hyperphosphorylation. These studies suggest that the growth of these human pancreatic cancer cells is still regulated by tumor-suppressor proteins.

Blotting, Western↗

PubMed1996

Progress toward hormonal therapy of gastrointestinal cancer.

OBJECTIVE: The authors discuss ongoing research to determine the mechanisms by which peptide hormones regulate growth of gastrointestinal cancer and ways in which this information might be used to develop noncytotoxic therapy. SUMMARY BACKGROUND: Approximately 100,000 people die of cancer of the gastrointestinal (GI) tract each year. Surgery is curative only when the tumor is localized. Chemotherapy and radiotherapy have limited efficacy for locally advanced or widespread disease. Various peptide hormones regulate the growth of these tumors. As for breast and prostate cancer, the growth regulatory effect of these hormones is being studied as a potential mechanism of hormonal therapy. CONCLUSION: Gut peptide hormones regulate the growth of GI malignancies through all-specific receptors and signal transduction pathways. Interruption or modification of these growth regulatory mechanisms may lead to specific noncytotoxic therapy. In combination with surgical extirpation, such hormonally based treatment may be curative of advanced GI cancer.

Animals↗

PubMed1996

All-trans-retinoic acid inhibits growth of human pancreatic cancer cell lines.

Retinoids are a class of molecules structurally related to vitamin A that have potent antiproliferative and differentiating effects on a variety of normal and neoplastic tissues. All-trans-retinoic acid (ATRA) has become a first-line chemotherapeutic agent in the treatment of certain leukemias; however, the effect of ATRA on pancreatic tumors is unknown. The purpose of this study was to determine the effect of ATRA on the growth characteristics of both exocrine and endocrine human pancreatic cancer cell lines. The in vitro growth of four cell lines was examined after treatment with a wide dose range of ATRA. The growth of all tumor cell lines was inhibited by ATRA in a dose-dependent fashion beginning at 0.1 microgram M. The in vivo growth of functioning human pancreatic carcinoid (BON) xenografts in Balb/c athymic mice was determined by treatment with several doses of ATRA over 1 month. The growth of BON tumors was inhibited in a dose-dependent fashion. These results suggest that ATRA exerts direct antiproliferative effects on both exocrine and endocrine human pancreatic cancers and may be useful in the chemotherapy of these tumors.

Animals↗

PubMed1996

Human monoclonal antibody against colon cancer.

The purpose of the study was to produce human monoclonal antibodies (hMcAb) against human colon cancer for use in radioimmunoimaging. Human-mouse heterohybridomas were developed by fusing SHM-D33 mouse-human hybrid heteromyeloma cells with human lymphocytes from colon cancer tumor-draining lymph nodes. The hybridomas capable of secreting human monoclonal antibodies were screened by using human colon cancer cell lines and pathological biopsies with ELISA and immunohistochemical methods. hMcAb clone H11 was selected for a large-scale antibody production, which was purified from mouse ascites. Biodistribution study demonstrated that specific uptake of 125I-hMcAb H11 by human colon cancer xenografts was significantly higher than by normal tissues. Radioimmunoimaging of human colon cancer xenografts exhibited distinct tumor visualization during the period of 72-96 hr after intraperitoneal injection of 125I-hMcAb H11. The development of human monoclonal antibodies such as hMcAb H11 may be useful for radioimmunodetection and therapy of colon cancer.

Animals↗

PubMed1995

Growth of mouse hepatocytes is stimulated by gastrin.

Hepatocyte growth is regulated by various growth factors, including epidermal growth factor (EGF) and insulin. Recently, several additional peptide hormones have been shown to stimulate growth of hepatocyte only in the presence of EGF or insulin and are thus termed secondary mitogens. Gastrin regulates growth of normal and neoplastic gastrointestinal tissues, but the effect on growth of hepatocyte is unknown. We examined the effect of gastrin on growth of a normal mouse hepatocyte (NMH) line established in our laboratory. Effect of gastrin-17 (G-17) (10(-8) to 10(-6) M) on growth of NHM cells was examined in either the presence or absence of EGF in the culture medium. Growth of NMH cells was evaluated by incorporation of either bromodeoxyuridine (BrdU) or 3H-thymidine and by counting cells. Presence of a cell-surface receptor for G-17 was determined by Scatchard analysis using 125I-G-17. In the presence of EGF, gastrin stimulated growth of NMH cells; in the absence of EGF, gastrin did not affect growth. The stimulatory effect of gastrin on NMH cells was blocked by JMV 320, a CCK-B type receptor antagonist. NMH cells possess a single, high affinity binding site for gastrin (Kd = 1.2 nM); EGF increased the gastrin binding capacity compared to non-treated cells (3.5 +/- 0.4 vs. 2.2 +/- 0.6 fmol/10(6) cells). G-17 stimulated growth of NMH cells through a single high affinity receptor for G-17 which pharmcologically appears to be the CCK-B type only in the presence of EGF and thus can be considered a secondary mitogen.

Animals↗

PubMed1995

Secretin potentiates cholecystokinin-stimulated amylase release by AR4-2J cells via a stimulation of phospholipase C.

Alterations in the activity of phospholipase C (PLC) are thought to be the primary intracellular events leading to pancreatic acinar cell exocytosis of zymogen granules. When multiple hormones, each of which may stimulate different signal transduction pathways, bind to cell surface receptors, the cell must integrate these signals into a common response through communication (cross-talk) among intracellular second messengers. We show that cholecystokinin (CCK) induces amylase secretion from AR4-2J pancreatic acinar cells via stimulation of PLC activity. Secretin indirectly stimulated the PLC pathway through cross-talk of the activated cAMP pathway to potentiate the CCK-stimulated amylase secretion. Therefore, secretin potentiated the acinar cell secretory response to CCK by cAMP-mediated cross-talk with the PLC signal transduction pathway.

Amylases↗

PubMed1995

Nerve growth factor as a mitogen for a pancreatic carcinoid cell line.

Carcinoid tumors are a group of neuroendocrine neoplasms distributed widely throughout the body but most commonly occurring in the gut. These tumors retain many characteristics of their neural crest origin, including secretion of neuroactive peptides and responsiveness to neurotrophic substances. Nerve growth factor (NGF), a neurotrophic protein involved in maintenance and differentiation of peripheral sympathetic and sensory neurons, regulates growth of several neural tumor cells by inducing a differentiated phenotype and subsequent inhibition of cell growth rate. We examined the actions of NGF in a functioning human pancreatic carcinoid cell line (termed BON). NGF has no effect on the cytoarchitecture or constitutive secretion of bioamines in this carcinoid cell line. NGF, however, stimulates the in vitro cellular proliferation of BON cells. BON cells possess mRNA for the NGF receptors (p75LNGFR and p140trkA) and membrane-associated tyrosine kinase activity is increased in response to NGF. Both the mitogenic activity of NGF, as well as the receptor-linked tyrosine kinase activity, can be abrogated in BON cells by the trkA inhibitor K-252a and specific anti-NGF antibody. Our studies demonstrate that NGF is a mitogen for this carcinoid cell line without effect on cellular phenotype or cytoarchitecture. NGF may play a role in the development and progression of human carcinoid tumors.

Carbazoles↗

PubMed1995

Role of calcium in the regulation of ornithine decarboxylase enzyme activity in mouse colon cancer cells.

Activity of ornithine decarboxylase (ODC), one of the rate-limiting enzymes in the pathway of polyamine biosynthesis, is regulated by various factors. In this study, we examined the role of Ca2+ in the regulation of ODC enzyme activity in mouse colon cancer cells (MC-26). KCl, a membrane-depolarizing agent that opens the voltage-dependent Ca(2+)-channel to increase intracellular Ca2+, decreased serum-induced ODC enzyme activity in MC-26 cells in a dose-dependent, reversible fashion. Both verapamil and nifedipine, inhibitors of the L-type voltage-dependent Ca(2+)-channel, decreased serum-induced ODC enzyme activity. W-7, a calmodulin inhibitor, decreased ODC enzyme activity in a dose-dependent, reversible fashion while trifluoperazine, another calmodulin inhibitor, failed to affect ODC enzyme activity in MC-26 cells. Our findings indicate that intracellular Ca2+ participates in the regulatory mechanism of ODC enzyme activity in MC-26 cells, although the exact role of Ca2+ is still unclear.

Animals↗

PubMed1994

Growth-regulatory effect of gastrin on human colon cancer cell lines is determined by protein kinase a isoform content.

Cell growth is regulated by various peptide growth factors through receptor-linked multiple intracellular signal-transduction pathways, such as the cyclic adenosine monophosphate (cAMP) pathway. cAMP activates cAMP-dependent protein kinase A (PKA) either to stimulate or inhibit cell growth. The effect on growth is determined by the presence of two isoforms of the regulatory (R) subunit of PKA; activation of RI alpha-type PKA leads to stimulation of growth, activation of RII beta-type inhibits cell growth. We determined whether the effect of gastrin on the growth of human colon cancer cells is determined by cell-specific content of PKA. We utilized two human colon cancer cell lines: LoVo, growth of which is stimulated by gastrin, and HCT116, growth of which is inhibited by gastrin. Activation of both types of PKA with 8-Br-cAMP mimicked the regulation of growth by gastrin; preferential activation of RII beta-type PKA with 8-Cl-cAMP inhibited growth of both cell lines. LoVo cells possess the predominantly RI alpha isoform of PKA at the mRNA and protein level; HCT116 cells possess predominantly the RII beta-type PKA. The cAMP-mediated regulation of growth (either stimulatory or inhibitory) by gastrin on these human colon cancer cells was determined by the predominant isoform of PKA.

8-Bromo Cyclic Adenosine Monophosphate↗

PubMed1994

Gastrin stimulates growth of human colon cancer cells via a receptor other than CCK-A or CCK-B.

Two receptors for cholecystokinin (CCK) have been isolated which also bind gastrin: CCK-A type and CCK-B type, both are coupled to phospholipase C (PLC) activation. However, identification of the "true" gastrin receptor remains controversial. We determined which CCK receptor mediated the trophic effect of gastrin on human colon cancer cells (LoVo). LoVo cells lack mRNA for either CCK receptor by Northern hybridization. Gastrin stimulated cyclic AMP production, not PLC activity, in LoVo cells. The trophic effect was not blocked by receptor antagonists for CCK-A (L364,718) or CCK-B (L365,260). The gastrin receptor pharmacology on LoVo cells and the lack of appropriate transcripts suggest that gastrin stimulated growth of these cells by a receptor other than CCK-A or CCK-B type and there likely exists another receptor for gastrin.

Cell Division↗

PubMed1994

Effects of gastrin on 3',5'-cyclic adenosine monophosphate, intracellular calcium, and phosphatidylinositol hydrolysis in human colon cancer cells.

Gastrin is a trophic factor for some human colon cancer cells. However, the signal-transduction pathways by which gastrin regulates growth are still unknown. We examined the effect of synthetic human gastrin-17 (G-17) on signal-transduction pathways and cell growth using 4 different human colon cancer cell lines (LoVo, COLO 320, HT-29, and HCT116). G-17 stimulated the production of cyclic AMP in LoVo, COLO 320, and HCT116 cells, while G-17 stimulated phosphatidylinositol hydrolysis and mobilization of intracellular calcium in HT-29 cells. The growth-regulatory effect of G-17 on these colon cancer cells (stimulatory on LoVo, COLO 320, and HT-29 cells; inhibitory on HCT116 cells) was well correlated with the effect of G-17 on the signal-transduction pathway in each cell line. We further examined the effect of a selective cholecystokinin-B type receptor antagonist, JMV 320, on G-17-induced signal-transduction pathways and G-17-regulated growth. In each cell line, the effect of JMV 320 on G-17-induced signal-transduction pathways was well correlated with that on G-17-regulated growth. G-17 appears to regulate, at least to some extent, growth of human colon cancer cells through gastrin receptor-linked signal-transduction pathways that are cell-specific.

Calcium↗

PubMed1994

Bombesin stimulates the in vitro growth of a human gastric cancer cell line.

Bombesin (BBS) and its mammalian equivalent, gastrin-releasing peptide (GRP), exhibit diverse biological functions, including that of a neurotransmitter, a regulator of gastrointestinal hormone release, and a trophic factor for various normal and neoplastic tissues. Bombesin stimulates the growth of normal cells of the stomach, pancreas, and bronchial epithelium as well as cells in breast cancer, gastrinoma, and small cell lung cancer. The purpose of this study was to determine whether BBS regulates the growth of a human gastric cancer cell line (SIIA) in vitro, and if so, to examine the mechanisms of signal-transduction that are involved. We found that BBS stimulated the growth of SIIA cells in vitro. The GRP receptor antagonists, BIM 26189 and BIM 26226, had no effect on growth of SIIA cells. Although these antagonists blocked the BBS-induced increase of [Ca2+]i, they failed to block the growth-stimulatory effect of BBS. BBS stimulated intracellular tyrosine phosphorylation of multiple proteins, with a predominant protein of apparent molecular weight of 125 kDa. Inhibition of intracellular tyrosine kinases by tyrphostin blocked the growth-stimulatory effect of BBS on SIIA cells. These results indicate that BBS exerts its trophic effect on SIIA cells through a receptor(s) linked to tyrosine kinase pathway, but not to the phospholipase C (PLC) pathway.

Adenocarcinoma↗

PubMed1994

Gastrointestinal hormones and cell proliferation.

There is no question that gut peptides are trophic for normal gut mucosa. Gut peptides can function in an endocrine, paracrine or autocrine fashion. We examined the effects of gut peptides on the growth of animal and human cancers of the gastrointestinal (GI) tract and pancreas in vivo and in vitro. We also examined the role of growth factors and bioamines in the regulation of growth of human endocrine tumors. Our studies have shown that gut peptides (gastrin, VIP, neurotensin, and bombesin) regulate growth of some cancers of the GI tract and pancreas. We have found that peptide action is mediated through specific receptors and that cell-specific differences in receptor expression occur. We have also begun to examine the intracellular signal-transduction pathways involved in endocrine and autocrine actions of these peptides on growth of GI cancers. We have found that cell-type-specific differences exist among the various signal-transduction pathways (cyclic AMP, phosphatidylinositol hydrolysis (PI), intracellular calcium ([Ca2+]i) mobilization, and tyrosine phosphorylation) and that different receptors for the same hormone may be linked to different signal-transduction pathways depending upon cell type. We have also found that autocrine growth regulation of human pancreatic carcinoid occurs through specific receptor-mediated signal-transduction pathways. We will discuss the mechanisms of action and potential therapeutic uses of manipulation of gut hormone levels or hormone antagonists to inhibit the growth of GI tract cancers.

Animals↗

PubMed1994

Cytocidal effect of high energy shock wave on tumour cells enhanced with larger dose and multiple exposures.

Cultured LLC-WRC256 (Walker rat carcinoma) cells were exposed to different doses of high energy shock waves (HESW). The immediate viabilities were 98% in the control cells, and 74%, 53% and 18% following 400, 800, and 1500 HESW treatment, respectively. Surviving cells in the 400 and 800-treated HESW demonstrated delayed upward growth rate curves, and the 1500 HESW-treated a downward curve. Agar clonogenic efficiencies for surviving cells were 36% (control), 20% (400 HESW), 15% (800 HESW) and 3% (1500 HESW). LLC-WRC256 tumours in Wistar rats were treated once every other day with 1500 HESW on a total of three occasions. Tumours treated with HESW grew more slowly (4.9 cm3) than those in the control (13.5 cm3). HESW fragmented cells and destroyed cell membranes and intracellular organelles. A histological examination of tumours treated with HESW demonstrated local haemorrhage with necrosis in the HESW focus area. Damage to the surrounding skin and soft tissue was slight and transient. These findings suggest that the growth of tumour cells can be suppressed in vitro and in vivo by treatment with HESW.

Animals↗

PubMed1994

Characterization of functional neurotensin receptors on human lymphocytes.

BACKGROUND: Neurotensin, a tridecapeptide, regulates gut motility, secretion, and mucosal growth; an immunomodulatory role for neurotensin has been postulated but not clearly defined. The purpose of this study was to determine whether neurotensin receptors (NTR) are present on peripheral blood lymphocytes (PBLs) and to characterize binding, functional, and molecular properties. METHODS: Iodine 125 labeled-neurotensin binding was determined for both human PBLs and the T-cell lines, Molt-4 and Jurkat, by Scatchard analysis. To analyze functional capacity of the NTR, PBLs were cultured in the presence of phytohemagglutinin (1 microgram/ml) with or without neurotensin and harvested at 48 hours after an 8-hour pulse of tritiated thymidine. For molecular analyses, RNA extracted from PBLs and T-cell lines was analyzed by either Northern hybridization with a labeled NTR cDNA or ribonuclease protection with an antisense human NTR probe. RESULTS: Scatchard analyses of neurotensin binding to human PBLs and MOLT-4 showed two classes of binding sites with different affinities. Incubation of phytohemagglutinin-stimulated PBLs with neurotensin significantly enhanced proliferation. Northern hybridization showed mRNA of the authentic NTR or a receptor subtype of close homology in PBLs and the T-cell lines; ribonuclease protection analysis identified the authentic human NTR in the MOLT-4 cell line. CONCLUSIONS: Using a combination of molecular techniques and Scatchard analysis, we have demonstrated the presence of a cell surface NTR with a high affinity for neurotensin on human PBLS and the MOLT-4 cell line. The functional role of NTR has been established by enhanced proliferation with addition of neurotensin. These data provide further evidence for a link between neurotensin and the immune system and suggest that neurotensin may play an important regulatory role in gut mucosal immune responses in vivo.

Blotting, Northern↗

PubMed1994

Experimental gene therapy of human colon cancer.

BACKGROUND: Gastrin regulates growth of human colon cancer cells by activation of the cyclic adenosine monophosphate (cAMP)-dependent protein kinase A (PKA). Gastrin and 8-Br-cAMP, a membrane-permeable cAMP analog, inhibit growth of HCT116 cells; both stimulate growth of LoVo cells. This dual effect on growth may be explained by relative amounts of the regulatory subunit (RI alpha or RII beta) of PKA within the cancer cells. Antisense oligodeoxynucleotides (ASO) to either RI alpha or RII beta inhibit protein translation of the target mRNA by sequence-specific binding; subsequently, cellular PKA content and the cAMP-mediated growth may be altered. We determined whether ASO to either the RI alpha or RII beta subunit altered the cAMP-mediated growth of HCT116 and LoVo human colon cancer cells. METHODS: HCT116 cells were treated with RII beta ASO (15 mumol/L, 4 days) and then treated with 8-Br-cAMP (25 mumol/L); tritiated thymidine incorporation was measured after 24 hours, and the cell number was determined on alternate days. Protein and mRNA levels of the RII beta subunit were determined by Western and Northern blotting, respectively. Similar studies with an ASO against the RI alpha subunit were performed on LoVo cells. RESULTS: RII beta ASO reversed the cAMP-mediated inhibition of growth of HCT116 cells, and RII beta ASO decreased the protein level of the RII beta subunit. RII beta ASO did not alter the basal growth of HCT116 cells. RI alpha ASO reversed the cAMP-mediated stimulation of growth of LoVo cells. CONCLUSIONS: The regulatory subunits of PKA are potential targets to alter growth of human colon cancer cells. Gene therapy directed to alter specific steps in signal transduction pathways may provide new therapeutic strategies.

8-Bromo Cyclic Adenosine Monophosphate↗