Cholecystokinin stimulates neuronal receptors to produce contraction of the canine colon

The motor effects of cholecystokinin 26-33-amide (CCK octapeptide; CCK-OP) and several purported CCK receptor antagonists on canine colonic circular muscle were determined in pentobarbital anesthetized dogs. Intravenous injections of CCK-OP had no effect on colonic motility at doses that contracted the gallbladder, stomach and duodenum. CCK-OP delivered by intraarterial injection to a small segment of the proximal colon produced a dose related increase in colonic motility with one-half maximum response at 12 ng/Kg and maximum response at 50 ng/Kg. The effects of intraarterial injections of several established CCK-receptor antagonists on proximal colonic responses to intraarterial injections of CCK-OP were determined. Proglumide, 10 mg/Kg, did not produce colonic contractions itself, but antagonized CCK-OP-induced responses. Carbobenzyloxy (CBZ)-CCK27-32-amide antagonized CCK-OP-induced colonic responses and also had no effect on basal colonic motility (0.1-1 and 5 micrograms/Kg). Neither compound antagonized acetylcholine- induced colonic responses. Butoxycarbonyl (BOC)-CCK31-33-amide increased basal colonic motility, but did not alter CCK-OP-induced responses at doses of 0.1 and 0.2 mg/Kg. Dibutyryl-cGMP at a dose of 0.1 mg/Kg did not affect basal motility or CCK-OP-induced contractions. At a dose of 1.0 mg/kg it increased basal colonic motility but did not affect CCK-OP-induced contractions. Pentagastrin increased colonic motor activity only at a dose of 5 micrograms/Kg, i.a., a much higher dose than effective doses of CCK-OP. The mechanism of CCK-OP-induced colonic motor effects also was determined. Atropine sulfate, 100 micrograms/Kg, i.v. significantly reduced both intraarterial acetylcholine-and CCK-OP-induced maximum colonic contractions. Tetrodotoxin, at intravenous doses that completely block neuronal activity, did not affect maximum acetylcholine-induced contractions but practically eliminated maximum CCK-OP-induced maximum colonic responses. In conclusion, intraarterial CCK-OP produces circular muscle contraction of the canine proximal colon that is mediated by stimulation of specific CCK receptors which produce the release of acetylcholine from cholinergic enteric neurons. Proglumide and CBZ-CCK27-32-amide are effective CCK receptor antagonists at these colonic neuronal receptors.     

青年和老年人结肠上皮的比较蛋白质组学研究

衰老的大肠上皮不仅多种生理功能下降,而且对大肠癌在内的多种衰老相关的肠道疾病易感性显著增加,但结肠上皮衰老及衰老的结肠上皮对癌症易感的分子机制仍然不清楚.为此,应用双向凝胶电泳(2-DE)技术分离青年人及老年人的正常结肠上皮的总蛋白质,图像分析识别差异表达的蛋白质点,基质辅助激光解吸电离飞行时间质谱(MALDI-TOF-MS)对差异表达的蛋白质点进行鉴定,免疫组化和实时定量(real-timequantitative)PCR检测部分差异蛋白,在青年人和老年人结肠上皮中的表达水平.得到了分辨率较高、重复性较好的青年人和老年人结肠上皮的2-DE图谱,质谱分析鉴定了17个结肠上皮衰老相关的蛋白质,免疫组化和real-timequantitativeRT-PCR证实了部分差异蛋白质的表达水平.研究结果提示,线粒体功能受伤、抗氧化能力下降是结肠上皮衰老的重要原因,4个差异蛋白质即guaninenucleotide-bindingproteinbetasubunit-likeprotein(Rack1)、stress-70protein、40SribosomalproteinSA和chlorideintracellularchannelprotein1可能与衰老的结肠上皮对癌症易感有关.     

Cholinergic stimulation enhances colonic motor activity, transit, and sensation in humans

The cholinesterase inhibitor neostigmine indirectly stimulates muscarinic M(1)/M(2)/M(3) receptors, thereby reducing colonic distension in acute colonic pseudo-obstruction. We investigated the dose-response profile for the colonic sensorimotor effects of neostigmine and bethanechol, a direct muscarinic M(2)/M(3) agonist in humans. A barostat-manometric assembly recorded phasic pressures, tone, and pressure-volume relationships (compliance) in the descending colon and rectum of 30 healthy subjects who received intravenous neostigmine (0.25, 0.75, or 1.5 mg; n = 15) or subcutaneous bethanechol (2.5, 5, or 10 mg; n = 15). Sensation to luminal distension was also assessed. Thereafter, the effects of neostigmine and bethanechol on colonic transit (geometric center) were compared with those of saline by scintigraphy in 21 subjects. Both drugs increased colonic phasic pressure activity, reduced rectal compliance, and enhanced urgency during rectal distension. Neostigmine also reduced colonic and rectal balloon volumes, reflecting increased tone by an average of 12% and 25% for the highest dose, respectively. Only neostigmine reduced colonic compliance, accelerated colonic transit [mean geometric center at 90 min 2.5 vs. 1.0 (placebo)], and increased pain perception during colonic distension. We conclude that neostigmine has more prominent colonic motor and sensory effects than bethanechol. Moreover, neostigmine induces coordinated colonic propulsion, perhaps by stimulating muscarinic M(1) receptors in the myenteric plexus.     

Heat shock protein 72 associated with CD44v6 in human colonic adenocarcinoma

CD44v6 is a splice variant of CD44 (CD44v), probably promoting cancer cell adherence to vascular endothelium and base membranes and enhancing the invasion and metastasis of colonic carcinomas. Heat shock protein 72 (HSP72) as a molecular chaperone has been confirmed to be overexpressed in epithelial carcinoma cells. There may be a possible association between the expression of HSP72 and CD44v6 during the growth and progression of colonic carcinoma cells. The aim of the study was to investigate the interaction between heat shock protein 72 and CD44v6 in human colonic carcinomas. The localization of HSP72 and CD44v6 in human colonic carcinomas was determined by immunohistochemistry and confocal laser microscopy. The interaction between HSP72 and CD44v6 in colonic carcinoma cells was analyzed by immunoprecipitation and Western immunoblots. Our results revealed that colonic carcinoma synchronously co-expressed higher levels of HSP72 and CD44v6 than that in adjacent normal colonic tissues. HSP72 and CD44v6 were mainly immunolocalized in the cytoplasm, and also immunolabelled on the cell membrane. Based on immunoprecipitation and Western immunoblots, we found that HSP72 was associated with CD44v6 precursor fragments in human colonic carcinoma cells. The interaction between HSP72 and CD44v6 in human colonic carcinoma cells may contribute to study the pathogenesis and immunotherapy of colonic carcinoma.     

The influence of adrenoceptor activity on cell proliferation in colonic crypt ipithelium and in colonic adenocarcinomata.

The effects of chemical sympathectomy and of the injection of amines or amine-receptor blocking drugs on cell proliferation in colonic crypts and in dimethylhydrazine-induced colonic carcinomata is examined in rats using a stathmokinetic technique. In animals which had been chemically sympathectomized by injection of 6-hydroxydopamine cell proliferation essentially ceased in the colonic crypts but continued at a normal rate in the tumours. Stimulation of alpha-adrenoceptors by metaraminol, a drug with properties similar to noradrenaline, caused acceleration of cell proliferation in colonic crypts but not in tumours. Conversely, blockade of alpha-adrenoceptors by phentolamine inhibited cell proliferation in crypts but not in tumours. Injection of adrenaline, predominantly a beta-adrenergic agonist, inhibited cell proliferation in the tumours but not in colonic crypts whereas blockade of beta-adrenoceptors by propranolol accelerated cell proliferation in tumours but not in colonic crypts. It is postulated that cell proliferation in the crypts of Lieberkühn in rat colon resembles that in rat jejunum in being controlled by the autonomic nervous system. However, tumour cell proliferation does not appear to be subject to such regulation.     

High body mass alters colonic sensory-motor function and transit in humans

There is increased prevalence of abdominal pain and diarrhea and decreased gastric sensation with increased body mass index (BMI). Our hypothesis is that increased BMI is associated with increased colonic motility and sensation. The study aim was to assess effect of BMI on colonic sensory and motor functions and transit. We used a database of colonic tone, compliance, and perception of distensions measured by intracolonic, barostat-controlled balloon, and gastrointestinal transit was measured by validated scintigraphy in healthy obese and nonobese subjects. Regression analysis was applied to assess the association of BMI with colonic sensory and motor functions. We included adjustments for sex differences, age, height, balloon volumes during distension, and psychological stress. Among 165 participants (87 women, 78 men), increased BMI was associated with decreased colonic compliance (P < 0.006, adjusted), decreased pain rating during distensions (P = 0.02, adjusted), and a higher threshold for pain (P = 0.042, adjusted). Sensation for gas, colonic tone, and contraction after meal ingestion were not significantly associated with BMI. Transit was assessed in 72 participants (41 women, 31 men); colonic transit was faster with BMI >30 kg/m(2) (P = 0.003 unadjusted, P = 0.08 adjusted for gender). In conclusion, BMI >25 kg/m(2) is associated with decreased colonic compliance and pain sensation; colonic transit is accelerated particularly with BMI >30 kg/m(2) in women. These data suggest that colonic dysfunction may contribute to diarrhea, but the cause of increased abdominal pain in obesity is not explained by the studies of colonic sensation and requires further study of afferent, spinal, and central mechanisms.     

Zinc L-carnosine protects colonic mucosal injury through induction of heat shock protein 72 and suppression of NF-kappaB activation

In this study, we investigated the effects of zinc L-carnosine, an anti-ulcer drug, on acetic acid-induced colonic mucosal injury and the correlation of these effects with expression of 72-kDa heat shock proteins (HSP72) and nuclear factor kappa B (NF-kappaB) activation in rat colonic mucosa in vivo. After intrarectal administration of zinc L-carnosine, the rats received intrarectal infusion of 5% acetic acid (1 ml). The colonic mucosal damage was evaluated by macroscopic assessments 24 h after the intrarectal infusion of acetic acid. Expression of HSP72 in rat colonic mucosa was evaluated by Western blot analysis before and after zinc L-carnosine administration. NF-kappaB activation was evaluated by electrophoretic mobility shift assays (EMSA). Zinc L-carnosine inhibited visible damage in rat colonic mucosa by acetic acid. Expression of HSP72 was significantly increased at 6 h after zinc L-carnosine administration. Furthermore, NF-kappaB activation in colonic mucosa was suppressed 6 h after zinc L-carnosine treatment. These results suggested that zinc L-carnosine protects the colonic mucosa against acetic acid by induction of HSP72 and suppression of NF-kappaB activation and zinc L-carnosine may be a novel therapeutic agent for the therapy of inflammatory bowel disease.     

Restraint stress stimulates colonic motility via central corticotropin-releasing factor and peripheral 5-HT3 receptors in conscious rats

Although restraint stress accelerates colonic transit via a central corticotropin-releasing factor (CRF), the precise mechanism still remains unclear. We tested the hypothesis that restraint stress and central CRF stimulate colonic motility and transit via a vagal pathway and 5-HT(3) receptors of the proximal colon in rats. (51)Cr was injected via the catheter positioned in the proximal colon to measure colonic transit. The rats were subjected to a restraint stress for 90 min or received intracisternal injection of CRF. Ninety minutes after the administration of (51)Cr, the entire colon was removed, and the geometric center (GC) was calculated. Four force transducers were sutured on the proximal, mid, and distal colon to record colonic motility. Restraint stress accelerated colonic transit (GC of 6.7 +/- 0.4, n=6) compared with nonrestraint controls (GC of 5.1 +/- 0.2, n=6). Intracisternal injection of CRF (1.0 microg) also accelerated colonic transit (GC of 7.0 +/- 0.2, n=6) compared with saline-injected group (GC of 4.6 +/- 0.5, n=6). Restraint stress-induced acceleration of colonic transit was reduced by perivagal capsaicin treatment. Intracisternal injection of CRF antagonists (10 microg astressin) abolished restraint stress-induced acceleration of colonic transit. Stimulated colonic transit and motility induced by restraint stress and CRF were significantly reduced by the intraluminal administration of 5-HT(3) antagonist ondansetron (5 x 10(-6) M; 1 ml) into the proximal colon. Restraint stress and intracisternal injection of CRF significantly increased the luminal content of 5-HT of the proximal colon. It is suggested that restraint stress stimulates colonic motility via central CRF and peripheral 5-HT(3) receptors in conscious rats.     

Short-chain fatty acids stimulate colonic transit via intraluminal 5-HT release in rats

We studied whether physiological concentration of short-chain fatty acids (SCFAs) affects colonic transit and colonic motility in conscious rats. Intraluminal administration of SCFAs (100-200 mM) into the proximal colon significantly accelerated colonic transit. The stimulatory effect of SCFAs on colonic transit was abolished by perivagal capsaicin treatment, atropine, hexamethonium, and vagotomy, but not by guanethidine. The stimulatory effect of SCFAs on colonic transit was also abolished by intraluminal pretreatment with lidocaine and a 5-hydroxytryptamine (HT)(3) receptor antagonist. Intraluminal administration of SCFAs provoked contractions at the proximal colon, which migrated to the mid- and distal colon. SCFAs caused a significant increase in the luminal concentration of 5-HT of the vascularly isolated and luminally perfused rat colon ex vivo. It is suggested that the release of 5-HT from enterochromaffin cells in response to SCFAs stimulates 5-HT(3) receptors located on the vagal sensory fibers. The sensory information is transferred to the vagal efferent and stimulates the release of acetylcholine from the colonic myenteric plexus, resulting in muscle contraction.     

The central effect of 5-hydroxytryptamine, gamma-amino-butyric acid and carbachol on thermoregulation in the neonatal guinea pig.

5-Hydroxytryptamine (10 microgram), gamma-amino-butyric acid (20 microgram) and carbachol (1 microgram) were injected through the soft skull into the lateral cerebral ventricle of guinea pigs aged 2 to 12 days, at the slightly subneutral ambient temperature of 30 degrees C. 5-Hydroxytryptamine (5-HT) produced no immediate change in oxygen consumption, whereas colonic temperature fell rapidly. One hour after injecting 5-HT, when colonic temperature had decreased by about 0.5 degrees C, oxygen consumption increased and after a lag of 30--40 min was followed by an increase in colonic temperature. Gamma-amino-butyric acid (GABA) increased oxygen consumption and colonic temperature after a latency of 30--60 min. Carbachol was followed by an immediate decrease in colonic temperature and oxygen consumption. Colonic temperature was below the pre-injection level throughout the five-hour period of observation.     

Supplemental silk protein, sericin, suppresses colon tumorigenesis in 1,2-dimethylhydrazine-treated mice by reducing oxidative stress and cell proliferation.

This study was done to discover the underlying mechanism of the inhibitory effect of sericin against colon tumorigenesis. Mice were fed a diet with 30 g/kg sericin for 115 d, and given a weekly injection of 1,2-dimethylhydrazine (10 mg/kg body weight) for the initial 10 wk. Dietary supplemental sericin caused a 62% reduction in the incidence of colonic adenoma (P<0.05), but did not affect the incidence of colonic adenocarcinoma. Sericin intake significantly reduced the number of colon adenomas. Consumption of sericin significantly reduced the BrdU labeling index of colonic proliferating cells and the expression of colonic c-myc and c-fos. The levels of colonic 8-hydroxydeoxyguanosine, 4-hydroxynonenal, and inducible nitric oxide synthase protein were significantly suppressed by sericin. The results suggest that dietary sericin suppresses the development of colon tumors by reducing oxidative stress, cell proliferation, and nitric oxide production.     

Dietary Iron Enhances Colonic Inflammation and IL-6/IL-11-Stat3 Signaling Promoting Colonic Tumor Development in Mice

Chronic intestinal inflammation and high dietary iron are associated with colorectal cancer development. The role of Stat3 activation in iron-induced colonic inflammation and tumorigenesis was investigated in a mouse model of inflammation-associated colorectal cancer. Mice, fed either an iron-supplemented or control diet, were treated with azoxymethane and dextran sodium sulfate (DSS). Intestinal inflammation and tumor development were assessed by endoscopy and histology, gene expression by real-time PCR, Stat3 phosphorylation by immunoblot, cytokines by ELISA and apoptosis by TUNEL assay. Colonic inflammation was more severe in mice fed an iron-supplemented compared with a control diet one week post-DSS treatment, with enhanced colonic IL-6 and IL-11 release and Stat3 phosphorylation. Both IL-6 and ferritin, the iron storage protein, co-localized with macrophages suggesting iron may act directly on IL-6 producing-macrophages. Iron increased DSS-induced colonic epithelial cell proliferation and apoptosis consistent with enhanced mucosal damage. DSS-treated mice developed anemia that was not alleviated by dietary iron supplementation. Six weeks post-DSS treatment, iron-supplemented mice developed more and larger colonic tumors compared with control mice. Intratumoral IL-6 and IL-11 expression increased in DSS-treated mice and IL-6, and possibly IL-11, were enhanced by dietary iron. Gene expression of iron importers, divalent metal transporter 1 and transferrin receptor 1, increased and iron exporter, ferroportin, decreased in colonic tumors suggesting increased iron uptake. Dietary iron and colonic inflammation synergistically activated colonic IL-6/IL-11-Stat3 signaling promoting tumorigenesis. Oral iron therapy may be detrimental in inflammatory bowel disease since it may exacerbate colonic inflammation and increase colorectal cancer risk.     

Central 5-hydroxytryptamine (5-HT) mediates colonic motility by hypothalamus oxytocin-colonic oxytocin receptor pathway

Gut-derived 5-hydroxytryptamine (5-HT) is well known for its role in mediating colonic motility function. However, it is not very clear whether brain-derived 5-HT is involved in the regulation of colonic motility. In this study, we used central 5-HT knockout (KO) mice to investigate whether brain-derived 5-HT mediates colonic motility, and if so, whether it involves oxytocin (OT) production in the hypothalamus and OT receptor in the colon. Colon transit time was prolonged in KO mice. The OT levels in the hypothalamus and serum were decreased significantly in the KO mice compared to wild-type (WT) controls. OT increased colonic smooth muscle contraction in both KO and WT mice, and the effects were blocked by OT receptor antagonist and tetrodotoxin but not by hexamethonium or atropine. Importantly, the OT-induced colonic smooth muscle contraction was decreased significantly in the KO mice relative to WT. The OT receptor expression of colon was detected in colonic myenteric plexus of mice. Central 5-HT is involved in the modulation of colonic motility which may modulate through its regulation of OT synthesis in the hypothalamus. Our results reveal a central 5-HT - hypothalamus OT - colonic OT receptor axis, providing a new target for the treatment of brain-gut dysfunction.     

Supplemental Silk Protein,Sericin, Suppresses Colon Tumorigenesis in 1,2-Dimethylhydrazine-Treated Mice by Reducing Oxidative Stress and Cell Proliferation

This study was done to discover the underlying mechanism of the inhibitory effect of sericin against colon tumorigenesis. Mice were fed a diet with 30 g/kg sericin for 115 d, and given a weekly injection of 1,2-dimethylhydrazine (10 mg/kg body weight) for the initial 10 wk. Dietary supplemental sericin caused a 62% reduction in the incidence of colonic adenoma (P<0.05), but did not affect the incidence of colonic adenocarcinoma. Sericin intake significantly reduced the number of colon adenomas. Consumption of sericin significantly reduced the BrdU labeling index of colonic proliferating cells and the expression of colonic c-myc and c-fos. The levels of colonic 8-hydroxydeoxyguanosine, 4-hydroxynonenal, and inducible nitric oxide synthase protein were significantly suppressed by sericin. The results suggest that dietary sericin suppresses the development of colon tumors by reducing oxidative stress, cell proliferation, and nitric oxide production.     

Binding of Shigella to rat and human intestinal mucin

Invasion of epithelial cells by Shigella is an early step in their pathogenesis. Adherence is generally presumed to be a prerequisite for invasion. This study examined the possibility of intestinal mucins serving as initial binding sites for clinical isolates of S. boydii and S. sonnei. The interactions of Shigella with rat and human small intestinal and colonic mucin were investigated. In solid phase binding assays, [³⁵S] labelled Shigella did not show any preferential binding to rat/human small intestinal mucin or to rat colonic mucin. On the other hand, Shigella bound specifically to human colonic mucin in a concentration-dependent manner. This specific binding to human colonic mucin was not by weak hydrophobic interactions and could not be attributed to the presence of contaminating glycolipids in the mucin preparation. The human colonic mucin receptor was sensitive to periodate treatment suggesting the involvement of the carbohydrate portion of the mucin. Reduction and alkylation of mucin enhanced adherence probably by exposing buried binding sites. The monosaccharides present in mucins were ineffective as hapten inhibitors as was the lectin wheat germ agglutinin suggesting that the mucin receptor is a more complex one. This study identifies, for the first time, the presence of a specific Shigella-binding site on the carbohydrate portion of human colonic mucin, which is not present in rat colonic mucin or in rat/human small intestinal mucin.     

Increased neurotensin receptor-1 expression during progression of colonic adenocarcinoma

The high affinity neurotensin receptor (NTSR1) mediates most of the biologic effects of neurotensin (NT), a 13-amino acid peptide that stimulates growth in certain cell types. NT is expressed in fetal but not differentiated colonic epithelium and is re-expressed in colonic adenocarcinoma. The cognate receptor, NTSR1, is also not expressed or is present at a low level in adult colonic epithelial cells but is expressed in most colon cancer cell lines. These observations suggest that altered NT-NTSR1 signaling may be associated with malignant transformation in the colon. To further understand the possible role of NTSR1 expression in colonic tumorigenesis and progression, we examined NTSR1 mRNA by in situ hybridization in normal colonic mucosa, adenomas, and colonic adenocarcinomas. NTSR1 mRNA expression was undetectable or weak in superficial differentiated epithelial cells of normal colonic epithelium, but adenomas and adenocarcinomas showed moderate to strong expression (p<0.05). Adenocarcinomas showed a higher level of expression compared to adenomas (p<0.05). Furthermore, adenocarcinomas that infiltrated into and beyond the muscularis propria showed a higher intensity of NTSR1 expression compared with tumors that were localized to the mucosa or submucosa. In some cases, infiltrating margins and foci of lymphovascular invasion showed a higher intensity of expression than the main mass of the tumor. These results suggest that increased NTSR1 expression may be an early event during colonic tumorigenesis and also contribute to tumor progression and aggressive behavior in colonic adenocarcinomas. NTSR1 may thus be a potential target for preventive or therapeutic strategies in colon cancer.     

Exogenous oxytocin reverses the decrease of colonic smooth muscle contraction in antenatal maternal hypoxia mice via ganglia

Oxytocin (OT) has been reported to have a potential protective effect on stress-induced functional gastrointestinal disorders. This study determined whether colonic contraction in adults was affected by antenatal maternal hypoxia, and whether OT is involved in antenatal maternal hypoxia induced colonic contraction disorder. Isometric spontaneous contractions were recorded in colonic longitudinal muscle strips in order to investigate colonic contractions and the effects of exogenous OT on the contraction in antenatal maternal hypoxia and control mice. Both high potassium and carbachol-induced contractions of proximal colon but not distal colon were reduced in antenatal maternal hypoxia mice. Exogenous OT decreased the contractions of proximal colonic smooth muscle strips in control mice, while it increased contractions in antenatal maternal hypoxia mice. OT increased the contractions of distal colonic smooth muscle strips in both antenatal maternal hypoxia and control mice. Hexamethonium blocked the OT-induced potentiation of proximal colon but not distal colon in antenatal maternal hypoxia mice. These results suggest that exogenous oxytocin reverses the decrease of proximal colonic smooth muscle contraction in antenatal maternal hypoxia mice via ganglia.     

Dietary Resistant Starch Alters the Characteristics of Colonic Mucosa and Exerts a Protective Effect on Trinitrobenzene Sulfonic Acid-induced Colitis in Rats

The protective effect of a dietary high-amylose cornstarch (HAS) against trinitrobenzene sulfonic acid (TNBS)-induced colitis was examined in rats. Rats were fed a HAS-free basal diet or, a 15% or 30% HAS supplemented diet for 10 d, and then received intracolonic TNBS to induce colitis and fed the respective diets for a further 8 d. HAS ingestion significantly protected colonic injuries as evidenced by lower colonic myeloperoxidase activity. Rats fed the HAS diet showed greater cecal short-chain fatty acid (SCFA) production than those fed the basal diet. Further, just before TNBS administration, HAS ingestion dose-dependently increased fecal and cecal mucin contents, and protein and nucleic acid contents in the colonic mucosa. HAS ingestion also reduced colonic permeability. The protective effect of HAS ingestion on TNBS-induced colitis is perhaps exerted through alterations in colonic mucosa, possibly due to cecal SCFA production.     

The major colonic cell mitogen extractable from colonic mucosa is an N terminally extended form of basic fibroblast growth factor.

Colonic growth factors (CGFs) were extracted from porcine intestinal epithelium and mucosa. Under acidic conditions, very little mitogenic activity (as assayed using murine 3T3 fibroblasts and a human colonic cell line) was extractable. However, by extracting at neutral or slightly alkaline pH, significant mitogenic activity for both the murine fibroblasts and human colonic carcinoma cell line could be detected. CGFs are present throughout the intestine and cecum. The epithelial mucosa of the distal colorectal region appeared to contain mitogens which were more potent for the colonic cells than the 3T3 fibroblasts. Purification of CGFs from the colonic mucosa required removal of associated mucin by pH precipitation prior to chromatographic fractionation. It was then possible to develop a complete purification (390,000-fold) scheme for the major CGF, an 18-kDa protein which bound to heparin-Sepharose. N-terminal sequence analysis yielded a single sequence (Q)SPGGAMAAGSITTLPALP, i.e. an N-terminally extended form of basic fibroblast growth factor. Apart from the substitution of Gly in bovine basic fibroblast growth factor by a Ser in porcine CGF, the proteins are identical. A similar extraction procedure using purified human colonic crypt epithelial cells yielded a mitogen for the human colonic cell line with similar chromatographic properties.