Is constipation caused by a loss of colonic interstitial cells of Cajal?

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particularly in the long term as demonstrated by a 5-year mortality of 49.5% versus 74.9% in the GVB and EVB groups, respectively. This difference in mortality is not fully understood, but one possibility is that the portal pressure gradient (PPG) may be a direct predictor of mortality as we have shown that it is significantly lower in the GVB group. Studies indicate a detrimental effect on mortality following TIPS insertion in patients with higher portal pressures, particularly shortly after the variceal bleed (Hepatology 1986;6:116 –117, Gastroenterology 1999;117:626 – 631, J Hepatol 1995;23:123–128). All of our patients had TIPS insertion using the same technique, although some patients with GVB were treated with thrombin injections. We have recently reported our experience using this endoscopic modality (Am J Gastroenterol 2002;97:1381–1385). Variceal hemorrhage particularly from gastric varices does occur frequently at PPG of ⱕ12 mm Hg, a finding we have reported in the past (Aliment Pharmacol Ther 1997;11:171–176, Am J Gastroenterol 1995;90:1994 –1996) and reinforced by the present study. This may be caused by the development of gastro-renal portosystemic shunts (Gastroenterology 1988;95:434 – 440), or the large size of the varices resulting in increased variceal wall tension (Semin Liver Dis 1986;6:318 –331). Most of the gastric varices in our study originated from the fundus or cardia and were of a large size. Other factors such as the presence of red spots, variceal size, and that of gastritis may be important, although it is noteworthy that the NIEC index only applies to esophageal varices (N Engl J Med 1988;319:983–989). Our findings highlight the need to reconsider the therapeutic targets for portal pressure reduction following TIPS. We suggest, particularly for patients with EVB, that a PPG post-TIPS of ⬍7 mm Hg is desirable to minimize variceal rebleeding. Large multicenter trials comparing TIPS with other endoscopic and pharmacological treatment modalities would be highly informative. In our experience, thrombin seems to be safe and effective, and therefore should be investigated further. In the absence of costeffective analysis, we would currently continue to recommend TIPS as first line treatment in patients with refractory GVB in the presence of a patent portal vein. DHIRAJ TRIPATHI, M.D., MRCP GEORGE THERAPONDOS, M.D., MRCP DORIS N. REDHEAD, M.D., FRCR PETER C. HAYES, M.D., Ph.D., FRCP

IS CONSTIPATION CAUSED BY A LOSS OF COLONIC INTERSTITIAL CELLS OF CAJAL? Lyford GL, He CL, Soffer E, Hull TL, Strong SA, Senagore AJ, Burgart LJ, Young-Fadok T, Szurszewski JH, Farrugia G (Mayo Clinic, Rochester, Minnesota; and Cleveland Clinic Foundation, Rochester, Minnesota). Pan-colonic decrease in interstitial cells of Cajal in patients with slow transit constipation. Gut 2002;51:496 –501. The pathogenesis of idiopathic slow transit constipation is unknown in most cases. Neuropathic and myopathic dysfunction and abnormal neuropeptide production have been described in selected patients. The interstitial cells of Cajal (ICCs) are increasingly recognized as the cell types that regulate gastrointestinal pacemaker function and coordinate neural input to gut smooth muscle. Furthermore, abnormal ICC populations have been described in a variety of clinical condi-

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tions with abnormal gut motor function. In the present investigation, Lyford et al. compared the distribution of ICCs in colonic tissues from patients with slow transit constipation undergoing subtotal colectomy with those from patients with resected colon cancers. Colonic specimens were acquired from 6 women with slow transit constipation defined by colonic transit values ⬎100 hours. All patients reported constipation for more than 10 years and failed stimulant and osmotic laxative therapies prior to referral for subtotal colectomy. Other causes of constipation were excluded by barium enema or colonoscopy to rule out structural abnormalities and by anorectal manometry and balloon expulsion testing to rule out pelvic floor dysfunction. Control samples were acquired from patients undergoing partial colectomy for colon cancer. Tissues from 12 patients (4 male, 8 female) were needed to provide 6 samples for each colon region. Frozen colonic tissue specimens were fixed, washed, and sectioned before staining for ICCs. Immunohistochemistry for ICCs was performed using c-kit polyclonal antibodies, and immunostained tissues were studied with laser scanning confocal microscopy. Specimens were divided into 4 anatomical layers: longitudinal muscle, myenteric plexus, circular muscle, and submucosal border. Results were compared in the cecum, right, transverse, and sigmoid colons. Three-dimensional software was used to quantify volumes occupied by ICCs in the entire thickness of the colonic muscularis and submucosa. Analyses were performed by investigators blinded to the patient identity or diagnosis. In control samples, ICCs were identified in all 4 anatomical layers with the highest volumes in the submucosal border and myenteric plexus (P ⬍ 0.05). ICC volumes did not significantly differ in different regions of the colon. In tissues from patients with slow transit constipation, ICC volumes were decreased by 40%–100% throughout (P ⬍ 0.05) with loss of cells in all anatomical layers. As in control specimens, ICC distributions were similar in different colonic regions in samples from constipated patients. The authors concluded that the decrease in ICCs is pancolonic in patients with slow transit constipation and speculated that the loss of this cell population may contribute to the pathogenesis of the condition. Comment. Idiopathic slow transit constipation is a heterogeneous disorder with many potential causes. Most often, the disorder is thought to result from visceral motor neuropathy. Histologic investigations of patients with slow transit constipation report decreases in colonic myenteric ganglia and neuron density (Histol Histopathol 1999;14:1119 –1134, Dis Colon Rect 2002;45:54 – 62). Additionally, myenteric neurotransmitters including vasoactive intestinal polypeptide, tachykinins, and enkephalins are reduced (Gastroenterology 1988;94:300 –310, Int J Colorect Dis 1998;13:208 –216), whereas nitric oxide levels may be increased (Dis Colon Rectum 2002;45:593– 600). Gut sensory abnormalities contribute to symptoms in some patients with slow colonic transit. Such individuals exhibit impaired perception of rectal distention as well as evidence of autonomic cholinergic nerve dysfunction (Scand J Gastroenterol 2001;36:32–38, J Auton Nerv Syst 1997;66:46 –52). Myopathic abnormalities described in slow transit constipation include decreases

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in circular muscle thickness and amphophilic inclusion bodies (Dis Colon Rect 1995;38:509 –513, J Pathol 2001;193:390 –397). Finally, factors external to the colon may participate in slow colonic transit. Impaired extrinsic serotonergic innervation with blunted gastrocolonic reflex activity has been described (Am J Physiol 2002; 283:G400 –G407). Colonic endocrine cells containing enteroglucagon and serotonin are decreased, whereas plasma hormone levels may be increased (cholecystokinin, pancreatic polypeptide) or decreased (peptide YY) (Scand J Gastroenterol 1999;34:1007–1011, Eur J Clin Invest 2000;30:709 –714). The ICCs are distinct from neurons and smooth muscle cells, and they play important roles in the regulation of colonic motility. Anatomic studies characterizing the distribution of ICCs measure immunoreactivity to c-kit, a proto-oncogene coding for a receptor tyrosine kinase. Such immunohistochemical studies identify at least 6 distinct ICC populations in the gut, including intramuscular ICC, ICC within the myenteric plexus, submucosal ICC in the colon, and ICC in the deep muscular plexus of the small intestine (Cell Tissue Res 1997;290:11–20). In contrast to the findings of the present investigation, others have reported regional variability in colonic ICC density with the highest numbers observed in the transverse colon (Am J Physiol 1998;275:G1309 –G1316). Evidence suggests that ICCs are important mediators of neurotransmission. ICCs in the myenteric plexus serve as a means of cellular communication between smooth muscle cells of the circular and longitudinal layers, exhibiting close appositions (Cell Tissue Res 1998;294:69 –79). In the small intestine, deep muscular ICCs are closely associated with excitatory tachykinin and inhibitory nitrergic neurons (Cell Tissue Res 1999;295:247–256). Similarly in the colon, intramuscular ICCs contact smooth muscle cells and nitrergic and cholinergic neurons (Cell Tissue Res 2000;302:331–342). In gastric tissues from mutant mice that lack intramuscular ICCs, cholinergic neural responses to electrical stimulation are profoundly reduced secondary to loss of synaptic contacts between the ICCs and motor neurons providing a functional importance for the histological observations (J Neurosci 2000;20:1393–1403). A second crucial function of ICCs is generation of the electrical pacemaker activities of the gastrointestinal tract. Intracellular recordings of colonic submucosal ICCs show continuous slow wave cycling (Am J Physiol 1989;257:C830 –C835). In colonic tissues devoid of ICCs, characteristic slow waves are absent (Am J Physiol 1991;260: G636 –G645). Intrauterine maturation of ICCs correlates with the initiation of electrical rhythmicity (Gastroenterology 1997;112:144 – 155). Furthermore, in mutant mice lacking ICCs, no spontaneous pacemaker activity is seen (Nature 1995;373:347–349, Am J Physiol 1996;271:G387–G399). Such loss of pacemaker function leads to disruption of organized luminal propagation (Gastroenterology 1998; 114:724 –736). In recent years, investigators have quantified ICC densities in human gastrointestinal diseases. Most studies have been performed in patients with disordered gut motility. ICCs are reduced in the colon in Chagas’ disease and megacolon, in the small intestine in megaduodenum, myopathic pseudo-obstruction, and long-standing diabetes, and in the pylorus in infantile hypertrophic pyloric stenosis (J Auton Nerv Syst 2000;80:108 –111, Gut 1999;45:775–779, Am J Gastroenterol 2002;97:2120 –2126, Am J Gastroenterol 1997;92:332–334, Gastroenterology 2001;121:427– 434, Gastroenterology 1996;111: 279 –288). ICC populations also are lost in illnesses not usually considered to be motility disorders including Crohn’s disease and ulcerative colitis, although these conditions exhibit altered motor activity as a consequence of the inflammatory infiltrates (Am J

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Gastroenterol 2002;97:118 –125, Gastroenterology 1996;111:1447– 1455). It is apparent that slow transit constipation, a more heterogeneous disorder than Chagas’ disease, pseudo-obstruction, or infantile pyloric stenosis, exhibits similar deficiencies in ICC distribution in the colon. In a prior study of patients with idiopathic constipation, ICC densities were decreased in the sigmoid colon compared with control samples (Gastroenterology 2000;118:14 –21). The present investigation by Lyford et al. expands on these findings, demonstrating uniform reductions in ICC density throughout the colon in this pancolonic condition. Furthermore, ICC populations in all layers of the colon are decreased, suggesting that cells responsible both for coordination of neurotransmission and for slow wave generation are impaired. In the discussion of their data, the authors postulate that this ICC defect may play an etiologic role in the generation of slow transit constipation. The findings of the present investigation do not offer insight into other issues related to the pathogenesis of slow transit constipation. No details regarding histologic abnormalities in the enteric ganglia or colonic smooth muscle in these particular patients are provided, thus it is uncertain if the ICC defect occurs in isolation or if it is part of a larger structural deficit in the cell types responsible for colonic propulsion. In a report of an infant born without an enteric nervous system, ICCs in the myenteric plexus were normal, although deep muscular plexus ICCs were degraded (Gastroenterology 2001;120: 561–567). Likewise, in another recent study of slow transit constipation, ICCs to most colon layers were reduced, but ICCs in the myenteric plexus were preserved (Int J Colorect Dis 2002;17:253– 258). In mice with hereditary aganglionosis, ICCs develop normally in regions without enteric neurons, indicating that different factors regulate development of the distinct cell types (Gastroenterology 1999;117:584 –594, Am J Physiol 2002;283:G445–G456). If the patients included in the present investigation exhibited generalized loss of ICCs and enteric neurons, this pattern might suggest that the histologic defect results from an undefined injury and that the ICC degradation per se is not the cause of slowed colonic transit. There is evidence from animal models to support the hypothesis that the ICC defect in slow transit constipation is a consequence rather than a cause of dysmotility. In mice, creation of a partial intestinal obstruction promotes disruption of ICC networks with loss of slow wave function orad to the blockage (J Physiol 2001;536:555– 568). This raises the possibility that the reduced ICCs observed in slow transit constipation are a consequence of chronic colonic retention with dilation. In this animal model, reversal of the intestinal obstruction leads to redevelopment of ICC networks with recovery of normal function. If the ICC defect in slow colonic transit is secondary to retained stool, perhaps early intervention with aggressive laxative therapy might prevent the histologic degradation observed in these advanced cases. These and other concerns indicate that much work needs to be done to confirm a pathogenic role for ICC defects in idiopathic constipation and other dysmotility syndromes. WILLIAM L. HASLER, M.D.

Reply. Ten years have passed since the initial reports that interstitial cells of Cajal (ICCs) generate the slow wave (J Physiol 1994;480:91– 97, Nature 1995;373:347–349). The discovery of a new molecule or of a cell type with a new function is usually followed by a marked increase in research in the area resulting in a series of reports that attempt to link the molecule or cell type to all diseases in the area of interest that do not yet have an etiology. This burst of activity is usually followed by a healthy skepticism and eventually steady state