ABSTRACT
BACKGROUND: Slow-transit constipation (STC) is recognized in children but the etiology is unknown. Abnormalities in substance P (SP), vasoactive intestinal peptide (VIP) and nitric oxide (NO) have been implicated. The density of nerve fibers in circular muscle containing these transmitters was examined in colon from children with STC and compared to other pediatric and adult samples. METHODS: Fluorescence immunohistochemistry using antibodies to NO synthase (NOS), VIP and SP was performed on colonic biopsies (transverse and sigmoid colon) from 33 adults with colorectal cancer, 11 children with normal colonic transit and anorectal retention (NAR) and 51 with chronic constipation and slow motility in the proximal colon (STC). The percentage area of nerve fibers in circular muscle containing each transmitter was quantified in confocal images. KEY RESULTS: In colon circular muscle, the percentage area of nerve fibers containing NOS > VIP > SP (6 : 2 : 1). Pediatric groups had a higher density of nerve fibers than adults. In pediatric samples, there were no regional differences in NOS and VIP, while SP nerve fiber density was higher in sigmoid than proximal colon. STC children had lower SP and VIP nerve fiber density in the proximal colon than NAR children. Twenty-three percent of STC children had low SP nerve fiber density. CONCLUSIONS & INFERENCES: There are age-related reductions in nerve fiber density in human colon circular muscle. NOS and VIP do not show regional variations, while SP nerve fiber density is higher in distal colon. 1/3 of pediatric STC patients have low SP or VIP nerve fiber density in proximal colon.
Subject(s)
Colon, Transverse/metabolism , Colon, Transverse/physiopathology , Constipation/physiopathology , Substance P/metabolism , Vasoactive Intestinal Peptide/metabolism , Adolescent , Adult , Age Factors , Animals , Biopsy , Child , Child, Preschool , Colon, Sigmoid/innervation , Colon, Sigmoid/metabolism , Colon, Sigmoid/physiopathology , Colon, Transverse/innervation , Female , Gastrointestinal Motility/physiology , Humans , Immunohistochemistry , Male , Nitric Oxide/metabolism , Nitric Oxide Synthase/metabolismABSTRACT
BACKGROUND: Studies in animals suggest that enteric neurons decrease in density or number with increasing age. Neurons containing nitric oxide (NO), vasoactive intestinal peptide (VIP) and Substance P (SP) have been implicated. In human large intestine, NO-utilizing neurons decrease during childhood or early adulthood but it is not known if the innervation of the muscle changes. This study examined the density of nerve fibres containing these transmitters in sigmoid colon circular muscle from children and adults. METHODS: Fluorescence immunohistochemistry using antibodies to neuronal NO synthase (nNOS), VIP and SP was performed on sigmoid colon from 18 adults with colorectal cancer, two children with familial adenomatous polyposis, and normal colon from nine children with Hirschsprung's disease. The percentage area of immunoreactive (IR) nerve fibres containing each transmitter in circular muscle was quantified in confocal images. KEY RESULTS: In the adult sigmoid colon circular muscle, the percentage area of nerve fibres containing nNOS>VIP>SP (6 : 2 : 1). Paediatric groups had significantly higher percentage area of nerve fibres containing nNOS, VIP or SP-IR than adults, with the decrease in nerve fibre density occurring from birth to 30 years. Circular muscle thickness increased between 12 and 30 years. Total nerve fibre area remained constant, while the muscle increased in thickness. CONCLUSIONS & INFERENCES: In human sigmoid colon circular muscle, there are reductions in nNOS-, VIP- and SP-IR nerve fibre density with growth from newborn to late adolescence but little further change with aging. The reduction in nerve density is due to an increase in circular muscle thickness rather than a loss of nerve fibres.
Subject(s)
Aging/physiology , Colon, Sigmoid/innervation , Muscle, Smooth/innervation , Nerve Fibers/metabolism , Adult , Age Factors , Aged , Aged, 80 and over , Cell Count , Child , Child, Preschool , Colon, Sigmoid/growth & development , Colon, Sigmoid/metabolism , Female , Humans , Immunohistochemistry , Male , Microscopy, Confocal , Middle Aged , Muscle Development/physiology , Muscle, Smooth/growth & development , Muscle, Smooth/metabolism , Nitric Oxide Synthase Type I/metabolism , Substance P/metabolism , Vasoactive Intestinal Peptide/metabolismABSTRACT
Evidence is presented that fibronectin is present on the platelet cell membrane and that it is a receptor for collagen in the platelet-collagen interaction. First, sodium dodecyl sulfate/acrylamide gel electrophoresis was performed on the proteins remaining attached to the surface of collagen after the removal of the remainder of the platelet by sonication. The material was relatively enriched in a glycoprotein that comigrated with cold-insoluble globulin (CIG), a form of fibronectin, and in other proteins which comigrated with myosin, actin, and tropomyosin. The presumptive presence of contractile proteins is consistent with the presence of microfibrillar proteins. Second, the collagen-attached material was shown to contain a protein that reacted with anti-CIG serum by immunoelectrophoresis. Third, when CIG was preincubated with fibrous collagen, the platelet-collagen interaction was inhibited. Fourth, rabbit anti-human CIG stimulated human platelets to secrete the contents of their dense granules. The stimulation was not due to antibody complexes present in the solution. Fifth, a protein was extracted from well-washed platelets and purified on affinity columns of anti-CIG antibodies. The isolated protein was found to bind to fibrous collagen.
Subject(s)
Blood Platelets/metabolism , Collagen/metabolism , Membrane Proteins/metabolism , Receptors, Drug/metabolism , Antigen-Antibody Complex , Antigens, Surface , Binding, Competitive , Cryoglobulins/immunology , Cryoglobulins/metabolism , Molecular Weight , Protein Binding , Serotonin/metabolismABSTRACT
The nature of the lipolytic activity released from chicken livers perfused with Krebs-Ringer buffer (pH 7.0) containing heparin (50 or 10 U/ml), fraction V albumin (3%), and glycerol (20%) was investigated. The nonrecirculating perfusates contained both previously described NaCl-resistant "liver lipase" as well as an apoLp-Gluactivated lipoprotein lipase (LPL). Crude perfusate lipolytic activity was separated on heparin-Sepharose columns into two enzymatic peaks which were eluted at mean NaCl molarities of 0.75 M (liver lipase) and 1.2 M (LPL). The liver LPL activity was stimulated 7-fold by human apoLp-Glu (half maximal activity at 1.5 microgram/ml) and inhibited by apoLp-Ala, apoLp-Ser, apoLp-GlnI, and apoLP-GlnII. Liver LPL was fully inhibited by anti-adipose LPL immunoglobulins. The "liver lipase" was not affected by apoLp-Glu (3-34 microgram/ml) or anti-adipose LPL immunoglobulins. The data demonstrate the presence in liver perfusates of a LPL with properties similar to adipose tissue lipoprotein lipase.
