403 Horm. Metab. Res. 7 (1975) 403-407
© Georg Thieme Verlag Stuttgart
Effect of Insulin on Human Intestinal Smooth Museie H.J. Arnqvist and B. Lundström Department of Pharmacology and Surgery, University of Linköping, Sweden
obesity. The effect of insulin (0.1 V/mi) on monosaccharide transport, amino acid transport, glucose metabolism and incorporation of amino acid into pro tein was examined. Also in human smooth muscle insulin stirnulated these parameters. The effects of insulin were moderate and appeared late. Material and Methods
Pieces of jejunum were obtained from 9 patients, 6 women and 3 men, undergoing shunt operations of the small intestine because of obesity. The operations were performed, afKey-Words: Human Smooth Muscle - Metabolism - Insuter an overnight fast, under general anesthesia and controlled lin respiration. The mean weight of the patients was 132 kg, the mean height 167 cm and the mean age 45 years. None of the patients had diabetes. Immediately after excision, the Introduction pieces of jejunum were placed in Krebs-Henseleit bicarbonate buffer at 370 C and brought to the laboratory within a couple The metabolie activity of vascular smooth muscle is of minutes. The mucosa was carefully removed with forceps probably an important factor in the development of and a pair of scissors. Histological control showed that no atherosclerotic diseases (Adams and Bayliss 1969, mucosa was present and that the preparation consisted only Wissler 1968, Wolinsky 1973). Since diabetes mellitus of the circular and longitudinal smooth muscle layer and the is associated with an increased frequency of vascular thin serosal layer which was not removed. No fat cells could diseases it is of interest to investigate how insulin in- be demonstrated in the muscJe preparation. After removal of the mucosa the muscJe layer was divided into pieces of fluences smooth muscle metabolism. sirnilar size weighing about 50 rng each.
In smooth muscJe preparations from anirnals moderate metabolie effects of insulin have been demonstrated in rat aorta (Wertheimer and Ben-Tor 1962), rabbit aorta (Mulcahy and Winegrad 1962), bovine mesenteric arteries and rabbit colon (Arnqvist 1971, 1972, 1973b, 1974). Monosaccharide transport, glucose uptake, glycogen synthesis, amino acid transport and incorporation of amino acid into protein were stirnulated by insulin in bovine mesenteric arteries and rabbit colon smooth muscJe. In comparison with rat diaghragm, which is an insulin sensitive and extensively used skeletal muscJe preparation, the effects of insulin on bovine mesenteric arte ries and rabbit colon smooth muscJe were qualitatively similar but quantitatively smaller and appeared later. It was therefore considered of interest to study if insulin influenced the metabolism of human smooth muscJe in a sirnilar way. In the present investigation, the effect of insulin on the metabolism of human intestinal smooth muscle was studied. The experiments were performed in vitra on the smooth muscJe layer of jejunum. Pieces of jejunum were obtained from patients undergoing shunt operations on the small intestine because of Received: 15 Jan. 1975
Accepted: 18 June 1975
The incubation procedure has been described in detail earlier (Arnqvist 1973b). In short, the tissue sampies were incubated at 37 0 C in 25 ml f1asks containing 4 ml Krebs-Henseleit bicarbonate buffer and a gas phase of 95% O 2 and 5% CO •. For determination of glucose uptake 10 ml f1asks containing 2.3 ml buffer were used. When added the concentration of insulin (monocomponent pork insulin from Novo) in the medium was 0.1 V/mI. The tissue accL:mulation of labe lied substrates was determined as previously described (Arnqvist 1974). After incubation in labelIed substrate the tissue sampies were rinsed in buffer for 10 s, bIotted, weighed and dissolved overnight in Soluene. The radioactivity of the tissue sampIes and of duplicate 100 fJl aliquots from the incubation media was counted in a liquid scintillation spectrometer (Packard Tri Carb). The accumulation of labelIed substrate was expressed as % distribution in total tissue (10 2 fJl/mg). Total tissue water was determined by drying to a constant weight at 100°e. Tissue sampIes for determination of glycogen content and incorporation of leueine into protein were rapidly frozen at -80 o C in Frigen (CFCI 3 ) containing solid CO 2 . The analytical methods used to determine glucose uptake, glycogen content and incorporation of leucine- 14 C into protein were the same as previously used (Arnqvist 1973a, 1974). In short, glucose was determined enzymatically by hexokinase and glucose-6-phosphate dehydrogenase (Slein 1962) and glycogen with an amyloglucosidase method (Keppler and Decker 1970). For determination of the incorporation of leucine- 14 C into pro tein the tissue sampies were homogenized in 10% trichloroacetic acid and the precipitated protein was purified accord-
Downloaded by: University of Pennsylvania Libraries. Copyrighted material.
Summary The action of insulin (0.1 V/mI) on the metabolism of human intestinal smooth muscJe was studied in vitro. The experiments were performed on the muscJe !ayer of human jejunum obtained from patients undergoing intestinal shunt operations because of obesity. Insulin significantly increased glucose uptake, glycogen content, the membrane transport of a-amino-isobutyric acid (AlB), the incorporation of leucine into protein and tended to increase the membrane transport of the nonutilizable model mono saccharide 3-0methylglucose. The effects of insulin were moderate and appeared after incubation times of 120 to 180 min.
H.J. Arnqvist and B. Lundström
ing to ArI'i11 and Ahren (1967). The dried protein was then weighed, dissolved in I ml Soluene and counted for radioactivity. Adjacent tissue sampies were used as test and control preparations. The significance of the effects was calculated by Student's t-test from the difference between these paired sampies. A difference resulting in p 0.05 was considered significant.
© Georg Thieme Verlag Stuttgart
Effect of Insulin on Human Intestinal Smooth Museie H.J. Arnqvist and B. Lundström Department of Pharmacology and Surgery, University of Linköping, Sweden
obesity. The effect of insulin (0.1 V/mi) on monosaccharide transport, amino acid transport, glucose metabolism and incorporation of amino acid into pro tein was examined. Also in human smooth muscle insulin stirnulated these parameters. The effects of insulin were moderate and appeared late. Material and Methods
Pieces of jejunum were obtained from 9 patients, 6 women and 3 men, undergoing shunt operations of the small intestine because of obesity. The operations were performed, afKey-Words: Human Smooth Muscle - Metabolism - Insuter an overnight fast, under general anesthesia and controlled lin respiration. The mean weight of the patients was 132 kg, the mean height 167 cm and the mean age 45 years. None of the patients had diabetes. Immediately after excision, the Introduction pieces of jejunum were placed in Krebs-Henseleit bicarbonate buffer at 370 C and brought to the laboratory within a couple The metabolie activity of vascular smooth muscle is of minutes. The mucosa was carefully removed with forceps probably an important factor in the development of and a pair of scissors. Histological control showed that no atherosclerotic diseases (Adams and Bayliss 1969, mucosa was present and that the preparation consisted only Wissler 1968, Wolinsky 1973). Since diabetes mellitus of the circular and longitudinal smooth muscle layer and the is associated with an increased frequency of vascular thin serosal layer which was not removed. No fat cells could diseases it is of interest to investigate how insulin in- be demonstrated in the muscJe preparation. After removal of the mucosa the muscJe layer was divided into pieces of fluences smooth muscle metabolism. sirnilar size weighing about 50 rng each.
In smooth muscJe preparations from anirnals moderate metabolie effects of insulin have been demonstrated in rat aorta (Wertheimer and Ben-Tor 1962), rabbit aorta (Mulcahy and Winegrad 1962), bovine mesenteric arteries and rabbit colon (Arnqvist 1971, 1972, 1973b, 1974). Monosaccharide transport, glucose uptake, glycogen synthesis, amino acid transport and incorporation of amino acid into protein were stirnulated by insulin in bovine mesenteric arteries and rabbit colon smooth muscJe. In comparison with rat diaghragm, which is an insulin sensitive and extensively used skeletal muscJe preparation, the effects of insulin on bovine mesenteric arte ries and rabbit colon smooth muscJe were qualitatively similar but quantitatively smaller and appeared later. It was therefore considered of interest to study if insulin influenced the metabolism of human smooth muscJe in a sirnilar way. In the present investigation, the effect of insulin on the metabolism of human intestinal smooth muscle was studied. The experiments were performed in vitra on the smooth muscJe layer of jejunum. Pieces of jejunum were obtained from patients undergoing shunt operations on the small intestine because of Received: 15 Jan. 1975
Accepted: 18 June 1975
The incubation procedure has been described in detail earlier (Arnqvist 1973b). In short, the tissue sampies were incubated at 37 0 C in 25 ml f1asks containing 4 ml Krebs-Henseleit bicarbonate buffer and a gas phase of 95% O 2 and 5% CO •. For determination of glucose uptake 10 ml f1asks containing 2.3 ml buffer were used. When added the concentration of insulin (monocomponent pork insulin from Novo) in the medium was 0.1 V/mI. The tissue accL:mulation of labe lied substrates was determined as previously described (Arnqvist 1974). After incubation in labelIed substrate the tissue sampies were rinsed in buffer for 10 s, bIotted, weighed and dissolved overnight in Soluene. The radioactivity of the tissue sampIes and of duplicate 100 fJl aliquots from the incubation media was counted in a liquid scintillation spectrometer (Packard Tri Carb). The accumulation of labelIed substrate was expressed as % distribution in total tissue (10 2 fJl/mg). Total tissue water was determined by drying to a constant weight at 100°e. Tissue sampIes for determination of glycogen content and incorporation of leueine into protein were rapidly frozen at -80 o C in Frigen (CFCI 3 ) containing solid CO 2 . The analytical methods used to determine glucose uptake, glycogen content and incorporation of leucine- 14 C into protein were the same as previously used (Arnqvist 1973a, 1974). In short, glucose was determined enzymatically by hexokinase and glucose-6-phosphate dehydrogenase (Slein 1962) and glycogen with an amyloglucosidase method (Keppler and Decker 1970). For determination of the incorporation of leucine- 14 C into pro tein the tissue sampies were homogenized in 10% trichloroacetic acid and the precipitated protein was purified accord-
Downloaded by: University of Pennsylvania Libraries. Copyrighted material.
Summary The action of insulin (0.1 V/mI) on the metabolism of human intestinal smooth muscJe was studied in vitro. The experiments were performed on the muscJe !ayer of human jejunum obtained from patients undergoing intestinal shunt operations because of obesity. Insulin significantly increased glucose uptake, glycogen content, the membrane transport of a-amino-isobutyric acid (AlB), the incorporation of leucine into protein and tended to increase the membrane transport of the nonutilizable model mono saccharide 3-0methylglucose. The effects of insulin were moderate and appeared after incubation times of 120 to 180 min.
H.J. Arnqvist and B. Lundström
ing to ArI'i11 and Ahren (1967). The dried protein was then weighed, dissolved in I ml Soluene and counted for radioactivity. Adjacent tissue sampies were used as test and control preparations. The significance of the effects was calculated by Student's t-test from the difference between these paired sampies. A difference resulting in p 0.05 was considered significant.
