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10.1245/ASO.2003.11.010
Annals of Surgical Oncology 10:801-809 (2003)
© 2003 Society of Surgical Oncology
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ORIGINAL ARTICLES

Decreased Expression of Stem Cell Factor in Esophageal and Gastric Mucosa After Esophagogastric Anastomosis for Cancer: Potential Relevance to Motility

Mario Nano, MD, FACS, Edda Battaglia, MD, Guido Gasparri, MD, FACS, Luca Dughera, MD, Pier Agostino Casalegno, MD, Graziella Bellone, PhD, Daniela Tibaudi, MD, Claudia Gramigni, PhD, Marco Ferronato, MD, Luigi Chiusa, MD, Monica Navino, MD, Mario Solej, MD, Marcello Dei Poli, MD and Giorgio Emanuelli, MD

From the General Surgery Section (MN, GG, MF, MS, MDP) and Internal Medicine Section (GB, DT, CG, MN, GE), Department of Clinical Physiopathology, S. Giovanni Battista Hospital of Torino (EB, LD, PAC); and Department of Biomedical Sciences and Human Oncology (LC), University of Torino, Torino, Italy.

Correspondence: Address correspondence and reprint requests to: Mario Nano, MD, FACS, Department of Clinical Physiopathology, University of Torino, Via Genova 3, 10126 Torino, Italy; Fax: 39-011-696-0170; E-mail: mario.nano{at}unito.it


   ABSTRACT
TOP
 ABSTRACT
INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
Background: Esophageal replacement with gastric tube is a well-established reconstruction of the alimentary tract after esophagectomy in cancer patients. The resulting molecular events in the transposed gastric tube and residual esophagus have yet to be investigated. Stem cell factor (SCF) was recently shown to be critical for signaling in gastrointestinal motility. SCF expression is here correlated with changes in mucosal morphology, acid and biliary reflux, and motility in the residual esophagus and gastric tube.

Methods: Thirteen patients surgically resected for squamous esophageal carcinoma with gastric tube replaced by esophagogastric anastomosis underwent upper endoscopy, esophageal manometry, 24-hour pH monitoring, and bile reflux detection. Esophageal and gastric mucosa samples were examined for SCF expression by immunohistochemical and semiquantitative reverse transcriptase-polymerase chain reaction analysis and for SCF serum levels by enzyme-linked immunosorbent assay.

Results: All patients showed severe residual esophagus hypoperistalsis and no gastric tube motor activity. The 24-hour pH monitoring was positive in most; 24-hour bile detection was mostly negative. SCF levels in the residual esophageal and gastric tube mucosa were dramatically decreased compared with those of normal subjects. The correlation between SCF and slow-wave activity was positive.

Conclusions: Hypomotility of the residual esophagus and gastric tube seems closely associated with disruption of the SCF/c-kit signaling pathway. However, the absence of notable relations between mucosal changes after chronic exposure to acid, biliary gastric content, and SCF expression indicates that this analysis cannot be considered part of endoscopic follow-up.

Key Words: Esophagogastric anastomosis • Stem cell factor • Motility • c-kit • Interstitial cells of Cajal


   INTRODUCTION
TOP
 ABSTRACT
 INTRODUCTION
MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
Esophageal replacement with a gastric tube is a well-established method for reconstruction of the alimentary tract after esophagectomy in cancer patients. The few studies on the pathophysiology of the intrathoracic stomach used as an esophageal substitute have reported that the gastric tube is almost inert and exposed to long biliary refluxes.1–4 However, data from two series of gastric replacements indicate the prevalence of endoscopic inflammation in the esophageal remnant in patients with long-term exposure.5 The pathogenetic mechanisms responsible for the mucosal lesions and symptoms often observed in these patients have not yet been elucidated.

The digestive and absorptive functions of the gastrointestinal (GI) tract depend on various mechanisms, mainly secretion and motility. Anatomical, functional, and electrophysiological investigations have shown that interstitial cells of Cajal (ICC) are critical for generation and propagation of slow-wave pacemaker activity and reception of regulatory inputs from the enteric nervous system.6 This unique class of stellate cells is located in the intramuscular space between the circular and longitudinal layers of muscularis propria in the stomach, small bowel, and colon, whereas they are distributed throughout the circular muscle layers and, in most areas, the longitudinal muscle layers in the esophagus, stomach, and colon.7

The expression of the type III tyrosine kinase receptor c-kit, the product of the proto-oncogene c-kit,8 is an established marker for ICC.9–11 c-kit activation by its natural ligand stem cell factor (SCF),12 also known as c-kit ligand (KL),13 mast cell growth factor,14 and steel factor,15 is essential for the development and maintenance of ICC networks.16 SCF, encoded at the Steel locus, is a growth factor that exists in soluble or transmembrane forms17 that play an important role in various biological phases, such as hematopoiesis, reproduction, and regeneration.18 In the GI tract, neuroblasts and smooth muscle cells express SCF and might provide proper signaling for the functional development and plasticity of ICC.19

Abnormal distribution and functioning of ICC might actually be involved in many disorders of GI transit, including achalasia, infantile hypertrophic pyloric stenosis, chronic intestinal pseudo-obstruction, congenital megacolon of piebaldism, Hirschsprung’s disease, inflammatory bowel diseases, slow-transit constipation, and some other disorders of GI motility.20 In addition, deregulated expression of the c-kit/SCF receptor/ligand system has been described in various tumors of hematopoietic, pigment cell, and germ cell origin,21–23 glioblastoma,24 breast carcinomas,25 small-cell lung cancer,26 GI stromal tumors,27 and colon carcinomas.28

In this study, we investigated the relationship between SCF expression and changes in mucosal morphology, acid and biliary reflux, and motility in the residual esophagus and the gastric intrathoracic tube after esophagectomy for cancer.


   MATERIALS AND METHODS
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
RESULTS
 DISCUSSION
 REFERENCES
 
Tissue Samples and Patients
A group of 13 patients was included in this study (all men; average age, 62 years; age range, 40–70 years). They had undergone surgical resection (by the same surgeon) at the General Surgery Section, Department of Clinical Physiopathology, University of Torino, Torino, Italy, between January 1989 and January 1996 for squamous carcinoma of the lower esophagus, which was replaced by an esophagogastric anastomosis (EGA) after construction of the gastric tube.29,30 Pyloroplasty was included in the gastric tube reconstruction and placed in the posterior mediastinum. The EGA was, in all cases, at the level of the azygos vein. All patients had been tumor free for 5 years. A second group of 20 subjects (13 men and 7 women; average age, 43 years; age range, 25–62 years) who were free from neoplastic or inflammatory diseases was used as control. The study was conducted in accordance with the ethical standards of the Committee for Human Experimentation of the San Giovanni Battista Hospital, Torino, and informed consent was obtained from each subject. All patients had undergone therapy with prokinetic and acid-suppressant drugs in the past, but therapy was suppressed 8 weeks before check-up. EGA patients and volunteers underwent upper endoscopy, and six biopsy samples were obtained from each patient and control subject. In EGA patients, two biopsy specimens were obtained from the esophageal mucosa 2 cm above the anastomosis, two from the midgastric tube, and two from the juxtapyloric gastric seat. In the volunteers, two biopsy specimens were obtained from the upper esophageal mucosa approximately 8 cm below the upper esophageal sphincter (UES), two from the midgastric seat, and two from the juxtapyloric seat. EGA patients underwent 24-hour pH monitoring and associated bile detection. Esophageal and gastric mucosa samples were frozen in liquid nitrogen immediately after removal and before messenger RNA (mRNA) extraction or were fixed in formalin and paraffin-embedded for immunohistochemical analysis. Serum samples from patients before and after surgery and from donors before endoscopy were stored at -70°C until use.

Twenty-Four–Hour pH Monitoring and Bile Detection
Twenty-four–hour esophageal and intragastric pH monitoring was performed with an antimony pH probe with three channels distanced 10 cm from one another, connected to a portable pH recorder (DGT4000; Medtronic, Northridge, CA). Recording sites were positioned, under fluoroscopic control, in the esophagus at 3 cm, in the midgastric tube at 13 cm, and in the juxtapyloric gastric seat at 23 cm below the UES, respectively.31 Data analysis was semiautomatic and was performed with Poligram software (Medtronic). During 24 hours of recording, the patient was asked to compile a diary with mealtimes, time spent in the supine position, and particular events, such as regurgitation, chest pain, or nausea/vomiting. In our system, the pH threshold value distinguishing acid reflux was 4; we considered the percentage of time with pH <4 as an expression of acid and pathologic reflux.32

The presence of bile in the esophageal-gastric lumen was studied concomitantly with two probes of the outpatient system Bilitec 200033 (Synetics Medical, Stockholm, Sweden); the probe transports the signal recorded in the stomach and esophagus to the electronic system. The probe is fiberoptic and flexible; its tip terminates with a polyvinyl chloride cap linked with a thin iron support. One probe was placed in the esophageal body at 3 cm below the UES, and a second probe was positioned 15 cm below the esophagogastrostomy, under fluoroscopy control. The patient was given a diary to note mealtimes and time spent in a supine position. The patient also had to avoid foods with a particularly dark color that could interfere with the optical system (for example, chocolate and soft drinks). Two light-emitting diodes (at 470 and 565 nm) were sources for a measurement and reference signal, respectively. A photodiode converts the light into electric signals that are processed, after amplification, by a microcomputer, which calculates the difference in adsorbence at 470 and 565 nm. This value is directly proportional to the bilirubin concentration in the sample studied and is therefore related to bile reflux.33

Esophageal Manometry
All patients underwent standard esophageal manometry with multichannel catheters perfused in polyvinyl chloride with an external diameter of 2.8 mm and eight channels (diameter, .8 mm) with orifices (diameter, .8 mm) 3 cm apart (Dentsleeve, Wayville, Australia) positioned with the proximal orifice 1 cm below the UES. Catheters were perfused with a pneumohydraulic pump (Andorfer like) with a constant perfusional water flux (.6 mL/min) for each channel.34 Each catheter was connected to an external transducer that transformed the pressure signal into an electric signal; this was then amplified and recorded by a recording system (Polygraf ID; Medtronic). Data analysis was semiautomatic. The test was performed with the patient supine. The manometric catheter was passed into the stomach and withdrawn to the proximal side hole with 1-cm steps. To evaluate esophageal body motility, patients performed at least 10 wet swallows with standard boluses (5 mL of water at 20°C–25°C) in different positions from 1 to 10 cm below the UES. The procedure was repeated after the catheter was repositioned 5 cm distally to record pressure changes over the entire length of the stomach.4 The manometric studies were compared with those from a group of healthy patients.

Histological Staining
Esophageal and gastric biopsy specimens, fixed in 10% neutral-buffered formalin and paraffin-embedded, were stained with hematoxylin and eosin and Giemsa stain for standard histological evaluation.

Immunohistochemical Detection of SCF
Formalin-fixed and paraffin-embedded biopsy specimens were stained with a standard avidin-biotin-peroxidase complex technique. Briefly, sections were deparaffinized and rehydrated in descending concentrations of alcohol. After blocking of endogenous peroxidase activity with 3% H2O2 in methanol and nonspecific binding sites with a protein blocker, a polyclonal goat anti-human SCF (Santa Cruz Biotechnologies, Santa Cruz, CA) diluted 1/100 was added. Controls included incubation of sections with non–immune-appropriate gamma globulins. Specificity of the antiserum against SCF was tested by preincubation with saturating amounts of recombinant SCF (R&D Systems, Minneapolis, MN), followed by immunostaining of the tissue sections. Staining patterns were evaluated using the immunoreacting score (IRS) proposed by Rammele and Stegner,35 in which IRS = SI x PP, where SI is staining intensity and PP is the percentage of positive cells. SI was determined as follows: 0, negative; 1, weak; 2, moderate; and 3, strong. PP was defined as follows: 0, negative; 1, 1% to 20% positive cells; 2, 21% to 50% positive cells; and 3, 51% to 100% positive cells. Ten visual fields from different areas of each specimen were chosen at random for IRS evaluation, and the average was calculated.

RNA Extraction and Semiquantitative Reverse Transcriptase-Polymerase Chain Reaction
Levels of SCF mRNA were assessed by semiquantitative reverse transcriptase (RT)-polymerase chain reaction (PCR) as described elsewhere.36 Briefly, total RNA was extracted from normal esophageal and gastric mucosa and from esophageal and gastric mucosa after EGA by using the single-step Trizol method (Life Technologies, Grand Island, NY). RNA (2 µg) was reverse-transcribed by Superscript II RT (Life Technologies). Each sample was subjected to an initial amplification with ß-actin–specific primers, as described elsewhere.28 On the basis of the amount of amplified ß-actin PCR product, an equal amount of reverse-transcribed product was amplified with the following SCF primer pairs37: 5'-CCATTGATGCCTTCAAGGAC-3' (sense) and 5'-GGCTGTCTCTTCTTCCAGTA-3' (antisense). The sequences of sense and antisense primers used for SCF were designed to detect both KL-1 (long form) and KL-2 (short form) mRNAs. PCR was performed in a reaction mixture containing 5 µL of complementary DNA, 200 µL of each deoxynucleotide triphosphate, .4 µM of each upstream- and downstream-specific primer, 1.5 µM of MgCl2, 2.5 U of Taq DNA polymerase (Life Technologies), and 1 µCi of [32P]deoxycytidine triphosphate (3000 Ci/mmol; DuPont/New England Nuclear, Boston, MA) in the reaction buffer supplied by the manufacturer. Thirty cycles were used: 4 minutes at 4°C for denaturation, 20 seconds at 58°C for annealing, and 30 seconds at 72°C for extension. The predicted sizes of SCF PCR products were 266 and 182 base pairs (bp). Samples were analyzed by electrophoresis through a 6% acrylamide Tris borate EDTA gel, followed by autoradiography and quantitation by Molecular Imager and Molecular Analyst software (Bio-Rad, Hercules, CA). Negative controls omitted RNA from complementary DNA synthesis and specific PCR amplification. Each experiment was repeated thrice.

Determination of SCF Concentration in Sera
SCF concentrations in sera were determined with an enzyme-linked immunosorbent assay kit (R&D Systems) by following the manufacturer’s instructions. All samples were evaluated twice.

Statistical Analysis
To compare values among patients and volunteers, the nonparametric Mann-Whitney test with Bonferroni’s significance-level correction was applied. The correlations between SCF expression and mucosal morphology, acid and biliary reflux, and motility were determined with nonparametric correlation Spearman’s R coefficient. The significance level was taken as P < .05.


   RESULTS
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
DISCUSSION
 REFERENCES
 
Histological and Functional Findings
Histological analysis of tissue samples from EGA patients gave the following results. For the esophagus, 1 (7.7%) of 13 patients showed epithelial parakeratosis, 2 patients (15.4%) showed mucosal hyperplasia, and 10 patients (76.9%) had normal histology. For the stomach, 5 (38.5%) of 13 patients showed chronic mild gastritis, 2 patients (15.5%) showed atrophic and chronically reactive gastritis, 3 patients (23%) showed fibrosis in the lamina propria, and 3 patients (23%) had normal histology. Gastric lesion patterns and distribution were random. In volunteers, endoscopy showed 1 case of mild gastritis and 1 case of esophagitis caused by acid reflux: these patients were excluded from the study, so the final number of control subjects was 18 rather than 20.

All patients showed severe hypoperistalsis of the residual esophagus (<30 mm Hg, mean amplitude of peristaltic waves). No motor activity, whether basal or evoked by the wet bolus, was found in the gastric tube. Twenty-four–hour pH monitoring determined a pathologic percentage of total time with pH <4 in the residual esophagus in 9 (69.2%) of 13 patients. In particular, in these nine patients, the mean percentage of the total time monitored (24 hours) during which the residual esophagus and gastric tube were exposed to pH <4 (either in the supine position or in orthostasis) was 29.7%, and the mean percentage of time at pH <4 when supine was 47.8%.

By our standard data,38 24-hour bile reflux monitoring was completely negative in 2 (15.4%) of 13 patients. In the remaining 11 patients with pathologic bile reflux, we found abnormal bile reflux in the gastric tube in 9 (81.1%) of 11 patients and abnormal bile reflux in the residual esophagus in 4 (36.4%) of 11 patients. No correlation was found between histological lesions in the esophagus and stomach, reflux score, and bile presence in any of the patients.

Immunohistochemical Detection of SCF in Normal Esophageal and Gastric Mucosa and in Esophageal and Gastric Mucosa After the EGA
Expression of SCF protein in the residual esophageal mucosa and in the gastric tube versus normal counterparts was determined by immunohistochemistry with specific polyclonal antiserum. Results in the form of a representative example are shown in Fig. 1A. In 60% of the operated patients, the gastric mucosa was negative for SCF with chronic inflammation in the mucosa; in the remainder, there was slight positivity, corresponding in 75% of cases to lesion-free gastric mucosa and in 25% of cases to slight chronic inflammation. Esophageal mucosa in operated patients was almost entirely negative to SCF, showing lesion-free mucosa in 60% of cases and slight hyperplasia of the epithelium in 40% of cases. In normal control subjects, the gastric mucosa showed intense cytoplasmic positivity on principal cells. Esophageal mucosa, on the contrary, showed diffuse positivity of the cytoplasmic type. To enable quantitative comparisons of staining patterns in different tissues to be made, immunoreactive scores were determined and evaluated by statistical analysis. As shown in Fig. 1B, statistically significant decreased levels of SCF in residual esophageal and gastric tube mucosa versus normal mucosa were found (P = .004 and P = .025, respectively).



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  		FIG. 1. (A) Representative immunohistochemical analysis of stem cell factor (SCF) protein expression in normal esophagus and stomach mucosa sections and in paired residual esophageal and gastric tube mucosa (original magnification x250). (B) Quantitative analysis of immunohistochemical staining of residual esophageal and gastric tube mucosa (n = 13) and normal counterparts (n = 18). The immunoreactive scores (IRS) were obtained as described in Materials and Methods. Results are expressed as mean ± SE.

 
Expression of SCF mRNA in Normal Esophageal and Gastric Mucosa and in Residual Esophageal and Gastric Tube Mucosa
We then determined the expression of SCF mRNAs by semiquantitative RT-PCR in paired residual esophageal mucosa and gastric tube biopsies from 8 of the 13 patients (for whom sufficient tissue was available) and in normal esophageal and gastric mucosa (n = 8). As shown in Fig. 2A, amplimers corresponding to the long (266 bp) and short form (182 bp) of SCF were generated from all tissue specimens analyzed.



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  		FIG. 2. (A) Analysis of stem cell factor (SCF) messenger RNA expression as detected by reverse transcriptase-polymerase chain reaction (PCR) in paired residual esophageal and gastric tube mucosa from 8 esophagogastric anastomosis (EGA) patients and in the normal counterparts (esophagus, lane 1; stomach, lane 2). A total of 266 and 182 base pairs (bp) SCF fragments, corresponding to the long and short forms, respectively, were generated using SCF-specific primers and DLD-1 cells as positive controls (lane 3). Lane 4 shows the lack of amplification products in the absence of a complementary DNA template. As quantitative and qualitative controls, ß-actin amplification products are shown in the bottom panel. (B) Densitometric values assigned to SCF (short and long form) bands were normalized for ß-actin expression, and the ratio of each SCF form to ß-actin was determined. Data are means ± SE (repeated 3 times). Statistically significant differences are indicated, KL, c-kit ligand.

 
To provide a more quantitative measure, densitometric values were assigned to SCF bands and normalized to ß-actin, and the ratios of each SCF form to ß-actin were determined. In agreement with immunohistochemical findings, a significant decrease in expression of both SCF forms was observed in gastric tube and residual esophageal mucosa versus normal counterparts (gastric tube vs. normal stomach: KL-1, P = .01; KL-2, P < .0001; residual esophagus vs. normal esophagus: KL-1, P = .0002; KL-2, P < .0001).

Correlation Between Motor Activity in Residual Esophagus and SCF Expression
Because GI motility depends on signaling via the SCF/c-kit tyrosine kinase pathway,9,39 we correlated the slow-wave activity of residual esophagus in patients after EGA with SCF expression. We found a positive correlation between SCF IRS and slow-wave activity (r = .79; P = .02).

SCF Levels in Sera of Patients and Normal Subjects
Because in certain human diseases serum SCF concentrations fluctuate,40 we measured soluble SCF levels in sera from esophageal and gastric cancer patients before and after resection versus normal controls by using a specific enzyme-linked immunosorbent assay. No statistically significant differences were observed between normal subjects (1043 ± 144 pg/mL) and patients (797.2 ± 221.2 pg/mL). In addition, no statistically significant differences in SCF serum levels were found in tumor patients before (797.2 ± 221.2 pg/mL) and after (843.7 ± 197.4 pg/mL) tumor resection.


   DISCUSSION
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
REFERENCES
 
The long-term functional results and clinical progress of intrathoracic stomach used as an esophageal replacement after esophagectomy in cancer patients have been widely studied, in some cases with opposing results.41–43 The molecular events occurring in the transposed gastric tube and residual esophagus as a result of this operative procedure have not yet been investigated. RT-PCR plus immunohistochemical analysis enabled us to determine, and to correlate with mucosa morphological changes, acid and biliary reflux, and motility, modulation in SCF expression in gastric tube and residual esophagus mucosa from a group of squamous esophageal carcinoma patients in whom the esophagus had been surgically resected and replaced with an EGA.

The study demonstrated that the gastric tube and, especially, the residual esophageal mucosa express significantly lower levels of SCF protein than their normal counterparts. Semiquantitative RT-PCR shows that normal specimens express both membrane-bound SCF long (KL-1) and short (KL-2) forms at the ratio of approximately 2.5:1. In gastric tube mucosa, KL-1 and KL-2 were decreased 3 and 8 times, respectively, whereas in residual esophageal mucosa, 5 and 8 times reductions were observed.

These findings are important in the light of previous studies showing that SCF, likely generated by circular smooth muscle layers in the GI tract, via c-kit receptor signaling, contributes to the development and maintenance of functionally competent ICC, the pacemaker cells responsible for enteric initiation of muscle slow-wave activity, propagation of electrical events, and modulation of neurotransmission.

Previous studies indicate that cell-associated SCF induces more persistent activation of c-kit kinase than the soluble form,44 generated by efficient proteolytic cleavage from KL-1. The alternative version of KL-1, KL-2, in which the major proteolytic cleavage site is removed by splicing, is a more stable membrane form of SCF and is thus more potent than KL-1 in activating the c-kit receptor. Thus, in gastric tube and residual esophageal mucosa, both SCF forms are dramatically decreased, in particular the more active one. This reduction is not associated with important pathologic changes in the mucosa. Esophageal epithelium from most of the patients (10 of 13) was normal, and gastric mucosa was likewise normal or presented signs of slight chronic gastritis. In contrast with results reported by other groups,43 we found chronic active gastritis in only two cases. Moreover, during this long survival time, no neoplastic or preneoplastic degeneration of the esophageal or gastric mucosa was observed. This discrepancy could be explained by the fact that our patients underwent surgery many years ago and had undergone prokinetic and acid-suppressor therapy for many years after surgery.

Acid secretion by the gastric tube and gastroesophageal reflux are joint determinant factors in the potential damage to the residual esophagus and intrathoracic stomach.45 We found a significant acid and biliary content in our patients, not only in almost all gastric tubes, but also in the residual esophagi. Despite vagotomy, acid secretion occurs; after a period of quiescence, hypergastrinemia decreases because the gastric tube is able to produce acid. However, the anatomical protective structures are no longer present, such as the lower esophageal sphincter and normal esophageal body motility, which is dramatically reduced in these patients. The gastric tube cannot be used for clearance, because it does not have coordinated oro-aboral motor activity.

In our patients, there was no correlation between morphological lesions and bile reflux, as reported by other groups; this may be a consequence of gastric stasis of solid meals for late tube emptying. A similar theory has been proposed by Clarke and Alexander-Williams46 in patients with gastric stasis after vagotomy, but without any kind of drainage. Despite demonstrations that vagotomy alone can cause mucosal changes, the etiology of gastritis in the gastric tube is uncertain.

Several reports claim that the gastric tube is inert and does not participate in the progression of food, which falls because of gravity.47–51 However, others claim that the gastric tube is a dynamic, contractile structure that recovers sufficient motor activity over the years.52–54 In addition, the gastric tube retains some secretory functions of both an exocrine and an endocrine nature.55 Our manometric results support the concept of an inert gastric tube, and all our patients showed true hypoperistalsis of the residual esophagus.

The most interesting finding was that, in patients with a great reduction of slow-wave activity in the residual esophagus and without any evidence of preneoplastic lesions or neoplastic recurrence, local SCF levels were particularly low. The cause of hypomotility is not entirely clear and is probably multifactorial. We can postulate that the defective expression of SCF, the critical factor for c-kit signaling in ICC,56 may contribute to the disruption of ICC networks, with concomitant functional losses in EGA patients in whom hypoperistalsis of the residual esophagus and the loss of motor activity of the gastric tube are observed. Studies showing that mice harboring inactivating mutations in either the SCF or c-kit genes lack ICC and that their GI tracts fail to display any slow-wave–type action potential57,58 are in favor of this hypothesis. It is difficult now to demonstrate the reason for the decreased production of SCF, although anatomical and functional changes caused by the gastroplasty might be involved. Therefore, with regard to the correlation between motility of the gastric tube and disruption of the ICC network, we believe that functional studies specifically targeted to this gastric portion are necessary, in parallel with the direct analysis of the SCF target cells, the ICC: in their seat (the muscular layer) on full-thickness visceral preparations, not on biopsy preparations that obviously include only the mucosa and submucosa. However, because it is difficult to perform animal studies that reproduce human conditions in full, the only possibility to verify the effect of gastroplasty on the functionality of the c-kit/SCF system would be to conduct a prospective study that evaluated SCF expression and immunolocalized cells of Cajal in full-thickness specimens taken before and after surgery from esophagus cancer patients. Ethical considerations would be difficult to reconcile with such a course.

The absence of a close relationship with mucosal changes after chronic exposure to acid and biliary gastric content suggests that, from this point of view, the c-kit/SCF system probably does not represent a useful prognostic marker in these patients, and its analysis cannot be considered part of the normal endoscopic follow-up of EGA patients. However, the finding that SCF levels are significantly reduced in residual esophageal and gastric tube mucosa from EGA patients may aid in the elucidation of the pathophysiological mechanisms responsible for their enteric motility disorders and provide a target for pharmacological treatment.

A new drug-delivery system using biodegradable microspheres has recently been developed to obtain sustained release of various drugs and mediators and successful targeting of specific organs.59 SCF delivery systems with these microspheres targeting the GI mucosa may be considered to be a therapeutic approach to improve gastric tube motor activity in EGA patients.

In summary, it may be hypothesized that the transposition of the stomach into the thorax, which involves an anatomical change of the gastric base area (which is physiologically involved in pacesetting the viscera and in which the ICC are particularly represented), brings about a functional remodeling caused by the lack of equilibrium produced in the ICC system.


   ACKNOWLEDGMENTS
 
The acknowledgments are available online at www.annalssurgicaloncology.org.

Supported by a research grant (692/27.001) from Regione Piemonte.


   FOOTNOTES
 
Recently stem cell factor (SCF) has been shown to be a critical factor for proper tyrosine kinase c-kit receptor signaling in gastrointestinal motility. This study aimed to correlate SCF expression and changes in mucosal morphology, acid and biliary reflux, and motility in residual esophagus and transposed gastric tube after reconstruction of the alimentary tract in esophagectomized cancer patients. The finding that hypomotility of the residual esophagus and gastric tube parallels defective expression of SCF suggests that disruption of the SCF/c-kit signaling pathway may contribute to the postoperative gastroesophageal functional loss often observed in these patients.

Received for publication November 14, 2002. Accepted for publication April 21, 2003.


   REFERENCES
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 

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