机械牵张对豚鼠膀胱组织Cajal间质细胞形态学及兴奋性的影响

目的 探讨机械牵张对豚鼠膀胱组织Cajal间质细胞(ICC)形态及兴奋性的影响.方法 建立雌性豚鼠膀胱颈部分梗阻(PBOO)模型作为实验组.并设假手术组作为对照.术后4周取膀胱组织制片,免疫荧光染色观察2组ICC形态和分布情况;胶原酶消化豚鼠膀胱制细胞悬液,免疫荧光标记,流式细胞仪检测2组c-kit阳性细胞比率.在弹性硅胶膜七原代培养膀胱ICC,利用Fluo-4钙荧光指示剂检测机械牵张对ICC钙信号的影响.结果 2组豚鼠膀胱组织铺片均可见c-kit染色阳性典型长梭形有突起的ICC,主要分布于平滑肌肌束间;PBOO组膀胱平滑肌间质增厚,c-kit染色阳性细胞数量及其突起明显增多,互相连接呈网络状;PBOO组c-kit阳性细胞比率为(6.7±1.7)%,显著高于对照组的(1.0±0.5)%,差异有统计学意义(P＜0.05).体外培养的ICC可检测到自发性钙波,机械牵张刺激可诱导ICC发生钙波增强现象.结论 机械牵张可诱导膀胱ICC兴奋性增强;PBOO膀胱组织中ICC数量及相互联系显著增多.ICC可能参与了膀胱牵张感受功能并在长期牵张应力负荷下发生一定的适应性改变.     

Interstitial cells of Cajal in the human normal urinary bladder and in the bladder of patients with megacystis-microcolon intestinal hypoperistalsis syndrome

OBJECTIVE: To investigate the distribution of c-kit-positive interstitial cells of Cajal (ICCs) in normal bladder and bladders from patients with megacystis-microcolon-intestinal peristalsis syndrome (MMIHS, a rare congenital and generally fatal cause of functional intestinal obstruction in the newborn), the most characteristic feature of which is abdominal distension caused by a distended unobstructed urinary bladder. PATIENTS AND METHODS: Full-thickness bladder specimens were obtained from four infants with MMIHS and four controls, and processed as paraffin-wax and frozen sections. Sections were assessed using single immunohistochemistry with monoclonal and polyclonal anti-c-kit antibodies. Anti-alpha-smooth muscle actin (SMA) antibody was used to investigate the contractile apparatus in smooth muscle cells of the urinary bladder. Specimens were examined using light and confocal scanning microscopy. RESULTS: There were many c-kit positive ICCs in the normal urinary bladder, appearing as small, long, bipolar cells with only two long and several short processes. In contrast, ICCs were absent in the MMIHS bladder. alpha-SMA immunoreactivity was lower in MMIHS urinary bladder than in control sections. CONCLUSION: This study shows for the first time the presence of c-kit-positive ICCs in the normal human urinary bladder. The lack of ICCs in the MMIHS bladder may contribute to the voiding dysfunction in this disease.     

慢传输型便秘患者结肠壁内Cajal细胞的形态学研究

目的　慢传输型便秘 (STC)患者乙状结肠壁内Cajal细胞 (ICC)的形态学研究。方法　全层铺片、冰冻切片的免疫细胞化学染色及透射电镜观察。结果　正常成人乙状结肠壁内ICC主要分布在环肌内侧面与粘膜下层之间 (ICC SM)、环肌层内 (ICC CM)、纵肌层内 (ICC LM)及肌间神经丛周围 (ICC MP)。STC乙状结肠壁内ICC的数量均较正常对照组减少 ,其中 ,ICC SM和ICC CM减少尤为显著 ,约减少 6 0 %。铺片显示ICC -MP不仅数量减少 ,且突起的分支亦减少 ,彼此间不能形成完整的细胞网络。电镜观察可见上述部位的Cajal细胞内溶酶体聚集、脂质沉积 ,ICC SM突起间的缝隙连接较小、数量减少。结论　本研究结果提示ICC的这些病理改变可能与STC的发生、发展有关 ,但是 ,ICC的减少是该病的原因还是继发性损害的结果仍有待进一步研究     

Three-dimensional morphology of gut innervation in total intestinal aganglionosis using whole-mount preparation

BACKGROUND: Total intestinal aganglionosis (TIA) is a rare form of Hirschsprung's disease (HD). The aim of this study was to examine the 3-dimensional morphology of the myentric plexus of the entire gastrointestinal tract in a newborn with total intestinal aganglionosis. METHODS: Whole-mount preparations were made of the entire gastrointestinal tract using NADPH-diaphorase histochemistry and c-kit (a marker of interstitial cells of Cajal) immunohistochemistry. RESULTS: Whole-mount preparations of the esophagus, stomach, and duodenum showed 3-dimensional morphology of the myenteric plexus forming a meshlike network of nerve fibers, connected to each other and to ganglia. There were large numbers of NADPH-diaphrase-positive nerve fibers between the muscle fibers in the circular muscle layer. In esophagus, stomach, and duodenum, c-kit-positive interstitial cells of Cajal (ICC) formed a 3-dimensional network between the two muscle layers and also were abundant within the circular muscle layer. In the jejunum, ileum, and colon, the myenteric plexus was absent and was replaced by hypertrophic nerve bundles that stained weakly with NADPH-diaphrase. Circular muscle layer completely lacked NADPH-diaphrase-positive nerve fibers. The c-kit-positive ICCs in the jejunum, ileum, and colon were sparse and localized mainly around the nerve trunks between the circular and longitudinal muscle layers. CONCLUSIONS: Whole-mount preparation is an elegant 3-dimensional technique in which the relationship of branching and interconnecting nerve fibers to each other and to muscle can be seen clearly. Absence of myenteric plexus, lack of nitrergic innervation, and depletion of interstitial cells of Cajal in the bowel wall throughout the small and large bowel contribute to the inability of the smooth muscle to relax, thereby causing lack of peristalsis in TIA.     

慢传输性便秘乙状结肠Cajal间质细胞分布的免疫组化观察

目的：Cajal间质细胞（interstitial cells of Cajal，ICC）对结肠动力有重要的调控作用。研究旨在初步观察ICC在STC乙状结肠的分布情况。方法：用ckit免疫组织化学技术观察了12例慢传输性便秘（slow transit constipation，STC）患者和8例对照组患者乙状结肠ICC形态和分布情况，并进行半定量分析。结果：ICC广泛分布于结肠肌层中，包括纵肌层（LM）、肌间丛（MP）、环肌层（CM）和黏膜下环肌表面（SMB）。ICC在形态上主要表现为双极细胞和多突起细胞两种类型。ICC—MP和ICC—CM相对多于ICC—SMB和ICC—LM。与对照组相比，STC患者各个区域ICC均明显减少（P〈0．01）。部分病例ICC—SMB几乎消失。结论：STC结肠ICC数量显著减少，但ICC减少是原发性的还是继发于某种其他损害仍有待探讨。     

Cajal-like cells in the human upper urinary tract

PURPOSE: Interstitial cells of Cajal (ICCs) have an important role in the regulation of gut motility as they are responsible for the slow wave activity of smooth muscle. It is still unknown if ICCs also occur in the human upper urinary tract. Since these cells express and are marked by the c-kit receptor CD117, we investigated its occurrence and distribution along the human upper urinary tract. MATERIALS AND METHODS: Tissues from 56 human ureters, spanning proximal, middle and distal ureter segments, were analyzed by indirect immunohistochemistry using the alkaline phosphatase-anti-alkaline phosphatase method and double labeling immunofluorescence on consecutive tissue sections. Several monoclonal and polyclonal antibodies to c-kit receptor were used in combination with various cell markers for histiocytic, mast cell, endothelial, epithelial, neuronal, smooth muscle and stem cell differentiation. RESULTS: The c-kit receptor was found in 3 cell types of the ureter and in round or spindle-shaped cells. Due to their antigenic profile the first one was revealed as mast cells occurring in all layers of the ureteral wall except the urothelium. In contrast, the population of spindle-shaped cells was only marked by c-kit receptor, thus, resembling ICCs. These ICC-like cells were found among the inner and outer smooth muscle layers, and in the lamina propria. They showed a slight decrease from proximal to distal ureteral segments. However, unlike intestinal ICCs their cytomorphology differed and some cells, representing the third group of c-kit receptor positive cells, were found within the urothelium. CONCLUSIONS: Our data demonstrate the presence of ICC-like cells and their ubiquitous distribution in the human ureter. The physiological importance and pathological significance of these findings must be evaluated by functional studies and investigations of certain pathological with urinary outflow disturbance conditions.     

Cajal间质细胞在先天性巨结肠和巨结肠同源病结肠中的分布研究

目的研究先天性巨结肠（Hirschsprung’s disease，HD）和巨结肠同源病（allied Hirschsprung’s disease，AHD）肠壁内Cajal间质细胞（interstitial cells of Cajal，ICCs）的分布状态，探讨HD和AHD的发病机制。方法选择确诊为HD和AHD的患者各20例，取巨结肠根治术吻合口远端的全层肠壁作为实验组，另取16例正常结肠标本作为对照组。用鼠抗人c—kit单克隆抗体（CD117）标记ICCs，Image Pro-Plus图像分析系统检测ICCs。结果对照组中大量ICCs分布在肌间神经丛周围和环纵肌层内，ICCs包绕神经丛周围，肌层间ICCs连续分布；HD组远端肠管中肌层间和各肌层内ICCs明显减少甚至缺如，与对照组比较差异有统计学意义，P〈0．01；AHD组远端肠管中神经丛大小不一，ICCs分布差异大，大多数神经丛区ICCs减少，环肌层内ICCs明显减少，与AHD组和对照组比较差异有统计学意义，P〈0.01。HD组远端结肠中ICCs减少比AHD组显著，差异有统计学意义，P〈0．01。结论HD、AHD病变肠管中除了神经节细胞的异常外，同时存在ICCs异常；ICCs在HD和AHD的分布不同可能与两者临床症状差异有关；肠管中ICCs数量可能与临床症状及预后有一定关系。     

Immunocolocalization of the heme oxygenase-2 and interstitial cells of Cajal in normal and aganglionic colon

Purpose: Interstitial cells of Cajal (ICCs) are pacemaker cells that play an important role in the control of gut motility. Carbon monoxide (CO) has been proposed as an endogenous messenger molecule between ICC and smooth muscle cells in the gastrointestinal tract (GIT). Heme oxygenase-2 (HO-2) is the main physiologic mechanism for generating CO in human cells. The aim of this study was to investigate the immunocolocalization of the HO-2 and ICCs in normal and aganglionic bowel of Hirschsprung's disease (HD). Methods: Full-thickness specimens were obtained from aganglionic colon during pull-through operation from 10 patients diagnosed as having HD. Normal control large bowel specimens were collected from 4 patients during bladder augmentation procedures. Double immunostaining was carried out using c-kit and HO-2 antibodies. Immunolocalization was detected by means of confocal laser scanning microscopy. Results: HO-2 immunoreactivity (IR) was found in many ICCs present around the myenteric plexus, within the longitudinal and circular muscle layers and at the innermost part of the circular muscle layer in normal colon. In the aganglionic colon there was absence of HO-2 IR in the sparsely found ICCs. In the transitional zone of HD bowel the colocalization of HO-2 IR and ICCs was much reduced compared with controls. Conclusions: The results of this study provide the first evidence for the presence of HO-2 immunoreactivity in the ICCs in normal human colon and absence of HO-2 immunoreactivity in sparsely appearing ICCs in the bowel of HD patients. The lack of HO-2 in the ICCs in the bowel of HD patients may result in impaired intracellular communication between ICCs and SMCs causing motility dysfunction. J Pediatr Surg 38:73-77.     

Absence of the interstitial cells of Cajal in a child with chronic pseudoobstruction

Absence or altered distribution of the interstitial cells of Cajal (ICCs) has been described in association with intestinal pseudoobstruction in adults. We report the first pediatric case with regional absence of ICCs in the distal small bowel and colon associated with intestinal pseudoobstruction. This report highlights that abnormalities of the ICCs in intestinal pseudoobstruction should be considered early in the diagnostic workup of children with intestinal pseudoobstruction.     

Gastrointestinal stromal tumors (GIST) of the stomach as a cause of upper gastrointestinal bleeding

Gastrointestinal stromal tumors (GIST) is the most common mesenchymal tumor of the gastrointestinal tract. GIST is currently defined as a gastrointestinal tract mesenchymal tumor containing spindle cells (less commonly epitheloid cells or rarely both) and showing CD 117 (c-kit protein) positivity in more than 95% of cases. Although they may arise throughout the gut, the commonest site are stomach (60-70%), small intestine (20-30%), colorectum (5%) and esophagus (up to 5%). Rarely, GISTs develop in the retroperitoneum, omentum or mesentery. GIST originates from the intestinal cell of Cajal (ICC). ICCs are located in and around the myenteric plexus and are thought to function as intestinal pacemaker cells. Historicaly, GIST were often misclassified as leiomyomas or leiomyosarcomas. Subsequently, it has been determined that GISTs have distinct ultrastructural features and immunophenotypical markers compared with smooth muscle and smooth muscle tumors. GIST predominantly occur in middle aged and older patients, with no significant difference in the sex incidence. Data from the recent population study suggest an incidence of about 10-22 cases per million persons per year. Clinical presentation of GIST varies widely, and depends on tumor size and location. GISTs that caused symptoms tended to be larger with an average size of 6cm versus 2cm for asymptomatic GISTs. Symptoms are most commonly related to mass effect or bleeding. GISTs can grow very large before producing symptoms. Commonest symptom of gastric GIST is manifest or occult bleeding. Abudant, life-threateting bleeding that require urgent surgery is rare. For patient with primary, localized, nonmetastatic GIST, complete surgical resection represents the only chance for cure. Lymhadenectomy is not necessary, because lymph node metastasis is very rare. The 5 year survival rate in patients with resected primary GISTs ranges from 48-65%. Conventional chemotherapy and radiation therapy is ineffective in the treatment of GIST. Imatinib mesilate (a tyrosine kinase inhibitor) was confirmed to be effective against metastatic or unresectable GISTs.     

Loss of CD117 (c-kit)- and CD34-positive ICC and associated CD34-positive fibroblasts defines a subpopulation of chronic intestinal pseudo-obstruction

Chronic idiopathic intestinal pseudo-obstruction is a syndrome in which symptoms of intestinal obstruction are present in the absence of mechanical obstruction. Lack of normal pacemaker activity, usually generated by the interstitial cells of Cajal (ICC), could account for the apparent obstruction. ICC are normally located around and between the myenteric plexus ganglia and within muscle and also in the deep muscular plexus of the small bowel and the submuscular plexus of the large intestine, just within the circular muscle. ICC can be demonstrated immunohistochemically with CD117 (c-kit) as well as with CD34, although this is less specific. CD34 also stains a population of fibroblasts that are intimately associated with ICC. To determine whether there is a relative deficiency of ICC and CD34-positive fibroblasts in patients with chronic idiopathic intestinal pseudo-obstruction, tissue from 30 patients of large intestine and eight patients with small intestine pseudo-obstruction was obtained. Controls (large intestinal specimens from 12 patients, small intestinal specimens from six patients) were chosen from resections for Crohn's disease and colorectal neoplasia, both with and without dilatation. Examination of pseudo-obstruction cases identified 10 patients (nine large intestinal and one small intestinal) in which both CD117 and CD34 were absent or severely reduced in all three of the examined areas. In contrast, the control cases, including those with preobstructive dilatation, showed relatively constant ICC staining. These results suggest that there is a proportion of pseudo-obstruction cases in which the ICC are markedly reduced. These results also demonstrate that, in these cases, loss of the kit immunoreactivity is correlated with the loss of CD34 staining: this indicates that both the ICC and the CD34-positive fibroblasts associated with the ICC are absent. These findings will allow surgical pathologists to identify this subpopulation of patients with CIIP using tissue obtained by laparoscopic biopsy of the muscularis propria or surgical resection.     

Loss of interstitial cells of Cajal and gap junction protein connexin 43 at the vesicoureteral junction in children with vesicoureteral reflux

PURPOSE: Symmetrical muscular contractions and unidirectional peristalsis are prerequisites for competent valve function at the ureterovesical junction. Interstitial cells of Cajal (ICCs) are pacemaker cells that create and coordinate peristaltic motility. We investigated ureteral endings immunohistochemically to elucidate the presence of c-kit positive ICCs as well as the occurrence of gap junction protein connexin 43 in children with vesicoureteral reflux (VUR) to identify a possible pathogenic factor of an insufficient antireflux mechanism. MATERIALS AND METHODS: Distal ureteral specimens were obtained from 27 children undergoing antireflux surgery. Routine histological paraffin embedded sections were immunostained detecting the c-kit proto-oncogene to study the presence of ICCs as well as connexin 43 positive cells to determine the gap junction density within the ureteral wall. Age matched nonrefluxing ureteral endings served as controls. All investigations were done using high power field magnification for semiquantitative analysis and statistically analyzed. RESULTS: ICCs were present in refluxing and nonrefluxing ureteral endings. Healthy individuals displayed significantly more ICCs than children with VUR. Connexin 43 immunoreactivity was significantly decreased in all refluxing ureteral specimens, whereas it was homogeneously distributed in normal controls. CONCLUSIONS: C-kit positive ICCs are found at the ureterovesical junction. Refluxing ureteral endings significantly lack these pacemaker cells, implying a malfunctioning valve mechanism permitting VUR. A substantial decrease in gap junctions in this region adversely affects intercellular signaling, aggravating coordinated peristalsis, which is essential for a competent anti-reflux mechanism.     

Identification of c-kit-positive cells in the human prostate: the interstitial cells of Cajal

The prostate exhibits electric activity in the form of slow waves (SWs) and action potentials (APs). As the interstitial cells of Cajal (ICCs) are considered the pacemaker cells which generate the electric waves, we investigated the hypothesis that the prostate contains ICC. Prostatic biopsies were obtained from 15 healthy volunteers (mean age 36 +/- 3.8 SD years). They were subjected to c-kit immunohistochemistry. Controls for the specificity of the antisera consisted of tissue incubated with normal rabbit serum substituted for the primary antiserum. C-kit-positive cells were identified as fusiform with dendritic processes. The cytoplasm was granular and the nucleus large and oval. Mast cells, also c-kit-positive, were round and lacked the dendritic processes. Immunoreactivity was absent in the negative controls. There were cells in the prostate with morphological and immunological phenotypes similar to ICCs of the gut. We predict an abnormal distribution of these cells in prostatic diseases. The study of the integrity of these cells may prove to be a useful investigative tool in the diagnosis of prostatic diseases and in the planning of an appropriate treatment.     

便秘二号方对慢传输型便秘大鼠Cajal间质细胞及C-KIT表达的影响

目的:观察便秘二号方对结肠慢传输型便秘大鼠模型的疗效,并检测其对大鼠模型结肠组织Cajal间质细胞及C-KIT表达的影响。方法:32只大鼠随机分为正常组、模型组、实验组、盐水对照组,每组8只。正常组给予正常饮食,其余3组采用大黄递增灌胃法制作慢传输型便秘大鼠模型。造模成功后,实验组予中药便秘二号方(方由当归、白术、黄芪、升麻、杏仁、枳实、肉苁蓉组成)灌胃,盐水对照组予生理盐水灌胃。30d后以活性炭灌胃法检测结肠传输功能,以免疫组化法观察结肠组织Cajal间质细胞的形态和数量,采用RT-PCR法观察C-KITmRNA表达。结果:炭末推进长度模型组明显少于正常组(P0.05),实验组明显高于盐水对照组(P0.05),推进率比较差异均有统计学意义(P0.01);实验组与盐水对照组相比,可见各肌层Cajal间质细胞数量增加,细胞形态恢复;实验组与盐水对照组的Cajal间质细胞个数比较,实验组高于对照组,差异有统计学意义(P0.01);模型组C-KIT表达阳性率明显低于正常组,实验组C-KIT表达阳性率明显高于盐水对照组。结论:中药便秘二号方可以增强慢传输型便秘大鼠的结肠蠕动功能,增加结肠组织中Cajal间质细胞的数量,改善其形态,促进C-KIT表达。     

Immunolocalization of the gap junction protein Connexin43 in the interstitial cells of Cajal in the normal and Hirschsprung's disease bowel

BACKGROUND: Interstitial cells of Cajal (ICC) are pacemaker cells between gastrointestinal smooth muscles; they generate spontaneous slow waves of the smooth muscle layers and mediate neurotransmission. The cellular network of ICC is connected by Gap junctions to each other and to the smooth muscle cells. Although there have been several studies reporting distribution of ICC in the normal bowel and pathological conditions such as Hirschsprung's disease, there is little information on the crucial role of Gap junctions in the intercellular communication in the gut musculature. The aim of this study was to investigate the immunolocalization of the Gap junction protein Connexin43 in the normal and Hirschsprung's disease (HD) bowel using whole-mount preparation technique and confocal laser scanning microscopy. METHODS: Full-thickness bowel specimens were collected at pull-through operation from 8 patients diagnosed as having HD. Normal control large bowel specimens were collected from 12 patients during bladder augmentation operation. Whole-mount preparation was performed on all specimens and double immunostaining was carried out using anti c-kit and antiConnexin43 antibodies. The immunolocalization was detected with the help of confocal laser scanning microscopy. RESULTS: Connexin43 immunoreactivity appeared in and between the c-kit-positive cells and along the smooth muscle fibers of the normal bowel and ganglionic part of HD bowel. In the aganglionic part of HD bowel there was no expression of Connexin43. In the transitional zone of HD the Connexin43 staining was weak and colocalized only in the processes of the c-kit-positive Cajal cells. CONCLUSIONS: Results of this study show for the first time that Gap junctional protein Connexin43 is present in the ICCs, which form a 3-dimensional network in the normal bowel wall. The lack of expression of Connexin43 in the aganglionic bowel and reduced expression in the transitional zone of HD suggest that the impaired intercellular communication between ICCs and smooth muscle cells may partly be responsible for the motility dysfunction in HD.     

Ontogeny of interstitial cells of Cajal in the human intestine.

BACKGROUND/PURPOSE: Interstitial cells of Cajal (ICC) recently have been identified as intestinal pacemaker cells. Abnormalities in ICC are increasingly recognized in a number of neonatal disorders such as infantile hypertrophic pyloric stenosis, Hirschsprung's disease, and transient intestinal pseudo-obstruction. The aim of this study was to determine the fetal and postnatal differentiation and development of ICC in the human gastrointestinal tract to aid interpretation of pathological specimens. METHODS: Specimens of human gastrointestinal tract from (1) fetuses (9 to 17 weeks' gestation; n = 12), (2) premature and full-term neonates with non-gut motility-related disorders, (age 26 to 59 weeks' gestation; n = 13), and (3) children (age 4 months to 13 years; n = 7) were immunohistochemically stained with antibodies to c-kit(a marker for ICC) and protein gene product 9.5 (PGP9.5, a marker for neural tissue). RESULTS: (1) C-kit-positive ICC were present throughout the gut in all specimens including those from the earliest gestational ages. C-kit and PGP9.5 immunoreactivities were present in different cell populations. (2) The distribution of ICC varied with gestational age and with region of the gut. (3) Maturation of ICC networks continues postnatally in a region-specific manner. CONCLUSIONS: ICC are present from an early stage in human gut development. Interpretation of apparent abnormalities in ICC distribution as being of pathological significance should be tempered by the knowledge that ICC networks continue to develop postnatally and that ICC development varies throughout the gut.     

Assessing interstitial cells of Cajal in slow transit constipation using CD117 is a useful diagnostic test

Slow transit constipation (STC) is a colonic motility disorder that is characterized by measurably delayed movement of stools through the colon. The pathophysiology of STC is unclear and both the interstitial cells of Cajal (ICC) and cells of the enteric nervous system are believed to play an important role. The aim of this study was to compare the number and distribution of ICC and cells of the enteric nervous system in patients with a control group by means of immunohistochemistry. Formalin-fixed paraffin-embedded colonic sections were obtained from 15 patients, aged between 23 and 52 (mean age=37 y), who underwent colectomy for STC. Forty-five cases of normal colon from age and sex-matched nonobstructive colorectal cancer patients were selected as controls. By using c-kit (CD117) and PGP 9.5 immunohistochemical studies, ICC and enteric neurofilaments were demonstrated, respectively. The number of cells were counted under 40 x high-power field (HPF) in 3 layers of the colonic muscularis propria, that is, the inner circular muscle layer, the myenteric plexus, and the outer longitudinal muscle layer in both test and control groups. The mean number of ICC and enteric neurofilaments were significantly reduced in all 3 layers of the muscularis propria from STC patients compared with controls. This reduction was most significant in the inner circular muscle layer (P<0.0001). A cutoff value of 7 ICC per HPF in the inner circular muscle layer can be used as a further confirmation to the clinical diagnosis of STC in resected specimens.     

Bombesin can minimize impairments of interstitial cells of Cajal induced by FK506 in small bowel transplantation

Purpose

Interstitial cells of Cajal (ICC) are known as intestinal pacemaker cells and express c-kit on their membrane. Previously, we reported that FK506 had neurotoxicity to enteric ganglia, and bombesin (BBS) preserved them against FK506. The aim of this study was to investigate whether ICC was impaired by FK506 and whether ICC was preserved by BBS against FK506.

Methods

Twelve rats underwent allogeneic SBTx heterotopically and were divided into 2 groups as follows: group A underwent SBTx with FK506 and group B with FK506/BBS. All rats were administered FK506 daily. Either BBS or normal saline was infused continuously from day 14 to 28. Analysis of ICC was performed immunohistochemically with c-kit. Interstitial cells of Cajal were evaluated by counting the number of c-kit-positive clusters in each graft.

Results

The expression of c-kit accumulated around 60% of PGP9.5-positive enteric ganglia. The number of c-kit-positive clusters in group A was 22.3 ± 5.5 clusters per cross section (C/CS) and that in group B was 36.3 ± 5.1 C/CS. Interstitial cells of Cajal were well preserved in group B. There was a significant difference between groups A and B (P <.001).

Conclusion

Interstitial cells of Cajal were impaired by FK506 in allografts, and BBS could minimize the impairment of ICC against FK506.     

Reduced insulin and IGF-I signaling, not hyperglycemia, underlies the diabetes-associated depletion of interstitial cells of Cajal in the murine stomach

Damage to interstitial cells of Cajal (ICC), pacemakers, and mediators of neuromuscular neurotransmission in the gastrointestinal tract contributes to the pathogenesis of diabetic gastroenteropathy in both patients and animal models. ICC depletion in diabetes may result from chronic hyperglycemia or lost/ineffective insulin signaling. Because independent control of insulin and glucose concentrations is difficult in chronic in vivo studies, we used long-term organotypic cultures to address this problem. Murine gastric muscles were cultured in normoglycemic or hyperglycemic basal media with or without insulin or IGF-I for 1-3 months, the time required for gastroparesis and ICC damage to develop in diabetic mice. ICC were assessed by c-Kit immunohistochemistry and quantitative analysis of c-kit expression. Electrical pacemaking was studied by intracellular recording of slow waves. ICC survived for at least 34 days in unsupplemented normoglycemic media, but their networks, c-kit expression, and slow waves were profoundly reduced after 68 days. These changes could be entirely prevented by insulin or IGF-I supplementation. ICC networks were completely resistant to hyperglycemia for at least 72 days. Thus, hyperglycemia is unlikely to be responsible for the diabetes-associated depletion of ICC. In contrast, maintenance of ICC requires insulin or IGF-I, which are reduced or ineffective in diabetes.