- Email: [email protected]
Expression of somatostatin receptor subtype mRNA in human colon cancer and normal mucosa
201 USE OF EXENDIN(9-39)AMIDE TO DEFINE THE IA! 171"OAND IA' I'TTRO ROLES OF GLP-l(736)AMIDE IN TIlE RIDGULATION OF INSULIN SECRETION IL~" Kulkarni, Z Wang, R Wang. DM Smith. MA Ghatei & SR Bloont Francis Frase~ Labs. Division o f Endoerinologl, & Metabolism, Dept o f ?eletabolic Medicine, Du Cane Road. London 1472 O.~'A~ UK.
Glucagon-like-peptide-l(7-36)amide (GLP-1) has been postulated to be the pfiman~ hormonal mediator of the entero-insular axis but evidence has been indirect. The discovery, of exendin(939)amide. a GLP receptor antagonist, allowed this to be investigated further in viva and in vitro. The in viva studies involved measurement of post-prandial release in trained conscious rats after a standard meal. After fasting from 17:00 the previous evening, male Wislar rats ( - 250g) were injected subcutaneously at 09:00 the nexa morning with either saline (n=20) or exendin(9-39)amide (0.5 nmol/kg, hE20). Blood glucose was significantly increased in the exendin treated group eomparod with the saline "treated group at 15 rain (9.9 ± 0.2 mmol.q vs 8.6 - 0.2 mmolq, p- :0.001) and at 45 rain (8.7 ± 0.19 retool/1 vs 7.5 -.+.0.1 mmol/L p. 0.001). The plasma insulin level was significantly decreased in the exendin treated group compared with the saline treated group at 15 rain (224 ± 32 pmol/1 vs 431 ± 21 pmol/I, p- O.001 ) and at 45 min (208 ± 20 pmol/I vs 349 ± 38 pmolq. p<0.01). The h~ vitro effect of GLP-1 was tested on the glycoraldehyde-sensitive rat clonal cell line RINm5F. The stimulator)., effect of 100 renal/1 of GLP-I was completely blocked by 1 ~tmol,q of exendin(9-39)amide (n~5, p. 9.001). Exendin(9-39)amide failed to block the stimulatou' effects of glucagon (1 nmol,q), secretin (10 nmol,q). VIP (100 renal/I) and GIP (100 nmol/1) (each n=3. p=NS). These results suggest exendin(9-39)amide is a specific antagonist of GLP-I and that gtucagon stimulates insulin secretion by acting on its o~a~ receptor mid not via GLP-I receptors. Taken together the~ studies suggest exendin(9-39)amide is a useful tool tu define the role of GLP-I and that GLP-1 eon'trihutes to the physiological relea~ of insulin in the post-praodial period.
EXPRESSION OF SOMATOSTATIN RECEPTOR SUBTYPE mPdqA IN HUMAN COLON CANCER AND NORMAL MUCOSA S.A.M. Laws, A.C. Gough, J.N.Primrose University Surgical Unit, Southampton General Hospital, Southampton SO9 4X'Y Somatostatin peptides (SS-14 and SS-28) are potent inhibitors of mucosal growth in the gastrointestinal tract and their stable analogues have anti-proliferative activity on gastrointestinal cancer in some systems. It is now known that the peptides act through 5 cell surface re.ccptors 0aSSTR1-5) which have differing alYanities for the analogues and linkage to intraeellular effector systems. We have therefore made a systematic study of the expression of the hSSTK subtype's mR_NA in colon cancer and matched muoosal samples by the reverse transeriptas¢ polymerase chain reaction ('RT-PCR) using sub-type specific primers, the products being resolved by agarose gel dectrophoresis. 18 turnour and mucosal pairs were collected, snap frozen and total RNA extracted. The hSSTR mRNA was semi quantified by comparison with a housekeeping gene (15actin). Results have indicated that over 60*/, of tumour mucosa pairs express at least one hSSTR subtype. Almost no tissues expressed hSSTR4. The SS28 preferring subtype, hSSTRS, was found on approximately 30*/, of all colonic specimens in low copy numbers, hSSTR1 & 2 were the most commonly expressed receptors with expression rates of approximatdy 35% e.ash, in tumour and mueosa. Five metastases have been studied and only one rex.eptor 0aSSTR3) has been found to be expressed in one specimen.
Glucagon-like-peptide-l(7-36)amide (GLP-1) has been postulated to be the pfiman~ hormonal mediator of the entero-insular axis but evidence has been indirect. The discovery, of exendin(939)amide. a GLP receptor antagonist, allowed this to be investigated further in viva and in vitro. The in viva studies involved measurement of post-prandial release in trained conscious rats after a standard meal. After fasting from 17:00 the previous evening, male Wislar rats ( - 250g) were injected subcutaneously at 09:00 the nexa morning with either saline (n=20) or exendin(9-39)amide (0.5 nmol/kg, hE20). Blood glucose was significantly increased in the exendin treated group eomparod with the saline "treated group at 15 rain (9.9 ± 0.2 mmol.q vs 8.6 - 0.2 mmolq, p- :0.001) and at 45 rain (8.7 ± 0.19 retool/1 vs 7.5 -.+.0.1 mmol/L p. 0.001). The plasma insulin level was significantly decreased in the exendin treated group compared with the saline treated group at 15 rain (224 ± 32 pmol/1 vs 431 ± 21 pmol/I, p- O.001 ) and at 45 min (208 ± 20 pmol/I vs 349 ± 38 pmolq. p<0.01). The h~ vitro effect of GLP-1 was tested on the glycoraldehyde-sensitive rat clonal cell line RINm5F. The stimulator)., effect of 100 renal/1 of GLP-I was completely blocked by 1 ~tmol,q of exendin(9-39)amide (n~5, p. 9.001). Exendin(9-39)amide failed to block the stimulatou' effects of glucagon (1 nmol,q), secretin (10 nmol,q). VIP (100 renal/I) and GIP (100 nmol/1) (each n=3. p=NS). These results suggest exendin(9-39)amide is a specific antagonist of GLP-I and that gtucagon stimulates insulin secretion by acting on its o~a~ receptor mid not via GLP-I receptors. Taken together the~ studies suggest exendin(9-39)amide is a useful tool tu define the role of GLP-I and that GLP-1 eon'trihutes to the physiological relea~ of insulin in the post-praodial period.
EXPRESSION OF SOMATOSTATIN RECEPTOR SUBTYPE mPdqA IN HUMAN COLON CANCER AND NORMAL MUCOSA S.A.M. Laws, A.C. Gough, J.N.Primrose University Surgical Unit, Southampton General Hospital, Southampton SO9 4X'Y Somatostatin peptides (SS-14 and SS-28) are potent inhibitors of mucosal growth in the gastrointestinal tract and their stable analogues have anti-proliferative activity on gastrointestinal cancer in some systems. It is now known that the peptides act through 5 cell surface re.ccptors 0aSSTR1-5) which have differing alYanities for the analogues and linkage to intraeellular effector systems. We have therefore made a systematic study of the expression of the hSSTK subtype's mR_NA in colon cancer and matched muoosal samples by the reverse transeriptas¢ polymerase chain reaction ('RT-PCR) using sub-type specific primers, the products being resolved by agarose gel dectrophoresis. 18 turnour and mucosal pairs were collected, snap frozen and total RNA extracted. The hSSTR mRNA was semi quantified by comparison with a housekeeping gene (15actin). Results have indicated that over 60*/, of tumour mucosa pairs express at least one hSSTR subtype. Almost no tissues expressed hSSTR4. The SS28 preferring subtype, hSSTRS, was found on approximately 30*/, of all colonic specimens in low copy numbers, hSSTR1 & 2 were the most commonly expressed receptors with expression rates of approximatdy 35% e.ash, in tumour and mueosa. Five metastases have been studied and only one rex.eptor 0aSSTR3) has been found to be expressed in one specimen.