Multitarget Stool mRNA Test for Detecting Colorectal Cancer Lesions Including Advanced Adenomas.

Current approved non-invasive screening methods for colorectal cancer (CRC) include FIT and DNA-FIT testing, but their efficacy for detecting precancerous lesions that are susceptible to progressing to CRC such as advanced adenomas (AA) remains limited, thus requiring further options to improve the detection of CRC lesions at earlier stages. One of these is host mRNA stool testing. The aims of the present study were to identify specific stool mRNA targets that can predict AA and to investigate their stability under a clinical-like setting. A panel of mRNA targets was tested on stool samples obtained from 102 patients including 78 CRC stage I-III and 24 AA as well as 32 healthy controls. Area under the receiver operating characteristic (ROC) curves were calculated to establish sensitivities and specificities for individual and combined targets. Stability experiments were performed on freshly obtained specimens. Six of the tested targets were found to be specifically increased in the stools of patients with CRC and three in the stools of both AA and CRC patients. After optimization for the choice of the 5 best markers for AA and CRC, ROC curve analysis revealed overall sensitivities of 75% and 89% for AA and CRC, respectively, for a ≥95% specificity, and up to 75% and 95% for AA and CRC, respectively, when combined with the FIT score. Targets were found to be stable in the stools up to 3 days at room temperature. In conclusion, these studies show that the detection of host mRNA in the stools is a valid approach for the screening of colorectal cancerous lesions at all stages and is applicable to a clinical-like setup.

MBD4 frameshift mutation caused by DNA mismatch repair deficiency enhances cytotoxicity by trifluridine, an active antitumor agent of TAS-102, in colorectal cancer cells.

BACKGROUNDS: Trifluridine is an active antitumor component of TAS-102 that resembles 5-fluorouracil. Although patients with advanced colorectal cancer (CRC) exhibiting a mismatch repair (MMR) deficiency reportedly do not benefit from 5-fluorouracil-based chemotherapy and we previously reported that truncated methyl-CpG binding domain protein 4 (MBD4) enhances 5-fluorouracil cytotoxicity in MMR-deficient CRC cells, little is known regarding the effect of MMR deficiency on trifluridine cytotoxicity in CRC. AIM: We investigated whether trifluridine induces cytotoxicity in a DNA MMR-dependent manner and evaluated how truncated MBD4 alters trifluridine cytotoxicity. METHODS: We utilized the human CRC cell lines HCT116 (hMLH1-deficient cells) and HCT116+ch3 (hMLH1-restored cells) and compared their sensitivities to trifluridine. And we established 5-fluorouracil-refractory hMLH1-deficient cells and analyzed trifluridine cytotoxicity. Finally, we established truncated MBD4 overexpressed CRC cell lines, and compared trifluridine sensitivity.Results: The sensitivities of HCT116 and HCT116+ch3 to trifluridine were comparable. 5-Fluorouracil-refractory hMLH1-deficient cells treated with trifluridine showed an equal or greater sensitivity than non-5-fluorouracil-refractory cells. Moreover, MBD4tru cells were more sensitive than the control cells to trifluridine.Conclusions: Trifluridine induces cytotoxicity independently of the DNA MMR status as well as under 5-fluorouracil-refractory conditions, and the MBD4 frameshift mutation enhances trifluridine cytotoxicity.

A Stool Multitarget mRNA Assay for the Detection of Colorectal Neoplasms.

Noninvasive screening methods for the detection of colorectal cancers (CRC) at curable stages rely on the identification of specific biomarkers. Our group has shown that mRNA stool assays represent a powerful and robust approach for the prediction of colorectal neoplasms. In this methodological chapter, we describe the procedures to isolate good quality stool RNA and the steps to evaluate the levels of specific host mRNA markers such as ITGA6, MYC, and GADD45B using TaqMan-based quantitative and droplet digital PCR approaches.

Droplet digital PCR for quantification of ITGA6 in a stool mRNA assay for the detection of colorectal cancers.

AIM: To investigate the use of droplet digital polymerase chain reaction (ddPCR) for detecting host mRNA markers in stools as a non-invasive test for colorectal cancer screening. METHODS: ddPCR and quantitative PCR were compared side by side for their performance in the detection of ITGA6 and ITGA6A transcripts in stool samples obtained from patients with various types of colorectal lesions (advanced adenomas and stage II-IV colorectal cancers) and control (patients displaying no pathological findings) using duplex TaqMan reactions for both methods. ITGA6 and ITGA6A were chosen for this proof-of-concept study based on their relative medium and low abundance in stool samples, respectively, as established in a previous study. RESULTS: We found that the ddPCR and qPCR methods performed equally well in this TaqMan duplex assay for the detection of ITGA6 and ITGA6A transcripts in stools of patients with colorectal lesions. For ITGA6, receiver operating characteristic (ROC) curve analysis showed comparable areas under the curve of 0.91 (P < 0.0001) and 0.89-0.90 (P < 0.0001) for the prediction of advanced adenomas and colorectal cancers, respectively. ITGA6A, which was detected at very low levels in control patients, was found to be significantly elevated (over 40 times) in stage II and III colorectal cancers (P < 0.0002). Comparison of the two sets of data revealed a strong correlation of the copy numbers obtained by ddPCR and qPCR for both ITGA6 and ITGA6A. CONCLUSION: We found that ITGA6 and ITGA6A detection in stools of patients with colorectal cancers with ddPCR is comparable to that of qPCR using TaqMan assays.

Production of truncated MBD4 protein by frameshift mutation in DNA mismatch repair-deficient cells enhances 5-fluorouracil sensitivity that is independent of hMLH1 status.

Methyl-CpG binding domain protein 4 (MBD4) is a DNA glycosylase that can remove 5-fluorodeoxyuracil from DNA as well as repair T:G or U:G mismatches. MBD4 is a target for frameshift mutation with DNA mismatch repair (MMR) deficiency, creating a truncated MBD4 protein (TruMBD4) that lacks its glycosylase domain. Here we show that TruMBD4 plays an important role for enhancing 5-fluorouracil (5FU) sensitivity in MMR-deficient colorectal cancer cells. We found biochemically that TruMBD4 binds to 5FU incorporated into DNA with higher affinity than MBD4. TruMBD4 reduced the 5FU affinity of the MMR recognition complexes that determined 5FU sensitivity by previous reports, suggesting other mechanisms might be operative to trigger cytotoxicity. To analyze overall 5FU sensitivity with TruMBD4, we established TruMBD4 overexpression in hMLH1-proficient or -deficient colorectal cancer cells followed by treatment with 5FU. 5FU-treated TruMBD4 cells demonstrated diminished growth characteristics compared to controls, independently of hMLH1 status. Flow cytometry revealed more 5FU-treated TruMBD4 cells in S phase than controls. We conclude that patients with MMR-deficient cancers, which show characteristic resistance to 5FU therapy, may be increased for 5FU sensitivity via secondary frameshift mutation of the base excision repair gene MBD4.

Use of integrin alpha 6 transcripts in a stool mRNA assay for the detection of colorectal cancers at curable stages.

OBJECTIVE: An important criterion for colorectal cancer (CRC) screening is the ability to detect lesions at a curable stage. In the present study, we have assessed the integrin α6 subunit transcript (ITGA6) as part of a stool assay for the detection of colorectal lesions. RESULTS: In comparison with control samples, ITGA6 levels were found to be significantly increased at all stages (P < 0.01). Receiver operating characteristic analysis revealed areas under the curve of 0.89 for the prediction of CRC with 81% sensitivity and 88% specificity and of 0.90 for the prediction of advanced adenomas (Ad) with 75% sensitivity and 88% specificity. The ITGA6A variant was also found to be increased relative to ITGA6 in stage II and III CRCs. Combining ITGA6 with other selected transcripts and/or immunochemical fecal occult blood test (iFOBT) results further increased sensitivity and specificity for the detection of colorectal lesions. PATIENTS AND METHODS: ITGA6 detection used alone and under various combinations including detection of other mRNA markers and iFOBT was assessed on stool samples obtained from 175 patients (91 CRCs, 24 Ad and 60 healthy controls). CONCLUSIONS: These data confirm the usefulness and reliability of an mRNA stool assay for the detection of colorectal lesions. The validation of additional candidate genes and their analysis in multiplex qPCR represents a powerful and robust approach that can be combined with iFOBT results to improve the detection of colorectal lesions.

Induction of murine TNBS colitis is strictly controlled by a modified method using continuous inhalation anesthesia with sevoflurane.

BACKGROUND: Trinitrobenzenesulfonic acid (TNBS)-induced colitis is one of the most widely used experimental colitis models. However, there is no standard procedure for inducing colitis by TNBS because it is difficult to achieve a uniform distribution of colitis. We have developed a modified method of murine TNBS-induced colitis that involves inhalation anesthesia with sevoflurane combined with both single and repeated TNBS administrations. AIMS: To compare the usefulness of our newly developed method for inducing murine TNBS-induced colitis with that of conventional intraperitoneal anesthesia. METHODS: TNBS in ethanol was administered to C57BL/6J mice held in an inverted vertical position either under continuous inhalation anesthesia with sevoflurane, in accordance with our newly developed method, or by intraperitoneal injection with 2.5 % avertin, in accordance with the conventional procedure. Body weight change, cytokine profile, and histological findings were examined during the course of colitis. RESULTS: The dispersion of anesthesia time, TNBS retention time, and nadir weight during the course of colitis was decreased using the newly developed method compared with the conventional procedure. Optimization of the modified TNBS-induced colitis, as evidenced by the predominant expression of Th1 and Th17 cytokines on day 7, was attained by the injection of 2.25 mg TNBS in 55 % ethanol. Regulation of the TNBS retention time using inhalation anesthesia with sevoflurane allowed strict control of the disease severity of TNBS-induced colitis. Using the modified method we were also able to develop a chronic TNBS-induced colitis model by repeated TNBS administration without excessive mortality of the mice. CONCLUSIONS: Our modified method for murine TNBS-induced colitis using continuous inhalation anesthesia with sevoflurane provides a better experimental colitis model following both single and repeated TNBS administrations.

SPG3A-linked hereditary spastic paraplegia associated with cerebral glucose hypometabolism.

SPG3A-linked hereditary spastic paraplegia (HSP) is a rare autosomal dominant motor disorder caused by a mutation in the SPG3A gene, and is characterized by progressive motor weakness and spasticity in the lower limbs, without any other neurological abnormalities. SPG3A-linked HSP caused by a R239C mutation has been reported to present a pure phenotype confined to impairment of the corticospinal tract. However, there is still a debate about the etiology of this motor deficit with regard to whether it is peripheral or central. We herein report two patients who were heterozygous for a R239C mutation in the SPG3A gene. Two middle-aged Japanese sisters had been suffering from a pure phenotype of HSP since their childhood. Both patients had a significant decrease in glucose metabolism in the frontal cortex medially and dorsolaterally in a [(18)F]-fluorodeoxyglucose (FDG) positron emission photography (PET) study and low scores on the Frontal Assessment Battery. A real-time PCR analysis in normal subjects showed the frontal cortex to be the major location where SPG3A mRNA is expressed. The present finding that the frontal glucose hypometabolism was associated with frontal cognitive impairment indicates that widespread neuropathology associated with mutations in the SPG3A gene may be present more centrally than previously assumed.

A distinct expression pattern of the long 3'-untranslated region dicer mRNA and its implications for posttranscriptional regulation in colorectal cancer.

OBJECTIVES: Reduced expression of Dicer is associated with global downregulation of microRNAs. Primary Dicer transcripts can be processed at two alternative polyadenylation sites, generating two pools of messenger RNAs (mRNAs) that carry either a long or a short 3'-untranslated regions (3'UTRs), that both encode the same Dicer protein. The short 3'UTR Dicer mRNA is not regulated by miR-103/107. The aim of this study was to investigate the expression of total Dicer mRNA, long 3'UTR Dicer mRNA, and miR-103 in colorectal cancer (CRC). METHODS: Paired tumor and normal mucosal specimens were obtained from 66 patients with CRC. Real-time reverse transcription PCR of long 3'UTR Dicer mRNA, total Dicer mRNA and miR-103 was carried out using the TaqMan Expression assay and the TaqMan MicroRNA assay. RESULTS: The median expression level of coding Dicer mRNA in the tumors was significantly lower than that in normal mucosa (P<0.001). There was no significant difference in expression levels of long 3'UTR Dicer mRNA between the tumors and the normal mucosa (P=0.90). The median expression ratio of long 3'UTR Dicer mRNA to total Dicer mRNA in tumors was significantly higher than in normal mucosa (P<0.001). There was no significant difference in expression levels of miR-103 between the tumors and normal mucosa (P=0.17). There was no significant correlation between clinicopathological findings, such as stage, tumor location, and histological grade and expression levels of total Dicer mRNA, long 3'UTR Dicer mRNA, or expression ratio of long 3'UTR Dicer mRNA to total Dicer mRNA. CONCLUSIONS: These results suggest that both transcriptional and posttranscriptional dysregulation of Dicer expression may be involved in colon carcinogenesis.

Successful treatment of ulcerative colitis complicated by Sweet's syndrome by corticosteroid therapy and leukocytapheresis.

Ulcerative colitis is occasionally complicated by dermatological disorders presenting as extra-intestinal manifestations, including erythema nodosum and pyoderma gangrenosum. Sweet's syndrome is considered to be a rare cutaneous disease in patients with ulcerative colitis. To date, only 17 cases of Sweet's syndrome complicating ulcerative colitis have been reported in the English literature. Here, we report a case of a 41-year-old male who had been suffering from ulcerative colitis for 20 years. He was admitted to hospital with hematochezia, diarrhea and fever, and painful erythematous nodules on the face and arms. Histological examination of skin biopsies showed inflammatory cell infiltration composed mainly of neutrophils without evidence of necrotizing vasculitis, and the condition was diagnosed as Sweet's syndrome. The patient was treated with prednisolone and leukocytapheresis and the erythematous nodules on the skin, as well as the abdominal symptoms and endoscopic findings of ulcerative colitis, immediately improved. In this paper we report on this case and review the literature concerning ulcerative colitis and Sweet's syndrome.

EPH-EPHRIN in human gastrointestinal cancers.

Ever since its discovery two decades ago, the erythropoietin-producing hepatoma (EPH)-EPHRIN system has been shown to play multifaceted roles in human gastroenterological cancer as well as neurodevelopment. Over-expression, amplification and point mutations have been found in many human cancers and many investigators have shown correlations between these up-regulations and tumor angiogenesis. Thus, the genes in this family are considered to be potential targets of cancer therapy. On the other hand, the down-regulation of some members as a result of epigenetic changes has also been reported in some cancers. Furthermore, the correlation between altered expressions and clinical prognosis seems to be inconclusive. A huge amount of protein-protein interaction studies on the EPH-EPHRIN system have provided a basic scheme for signal transductions, especially bi-directional signaling involving EPH-ERPHRIN molecules at the cell membrane. This information also provides a manipulative strategy for harnessing the actions of these molecules. In this review, we summarize the known alterations of EPH-EPHRIN genes in human tumors of the esophagus, stomach, colorectum, liver and pancreas and present the perspective that the EPH-EPHRIN system could be a potential target of cancer therapy.

Biological effects of mutant ceruloplasmin on hepcidin-mediated internalization of ferroportin.

Ceruloplasmin plays an essential role in cellular iron efflux by oxidizing ferrous iron exported from ferroportin. Ferroportin is posttranslationally regulated through internalization triggered by hepcidin binding. Aceruloplasminemia is an autosomal recessive disorder of iron homeostasis resulting from mutations in the ceruloplasmin gene. The present study investigated the biological effects of glycosylphosphatidylinositol (GPI)-linked ceruloplasmin on the hepcidin-mediated internalization of ferroportin. The prevention of hepcidin-mediated ferroportin internalization was observed in the glioma cells lines expressing endogenous ceruloplasmin as well as in the cells transfected with GPI-linked ceruloplasmin under low levels of hepcidin. A decrease in the extracellular ferrous iron by an iron chelator and incubation with purified ceruloplasmin in the culture medium prevented hepcidin-mediated ferroportin internalization, while the reconstitution of apo-ceruloplasmin was not able to prevent ferroportin internalization. The effect of ceruloplasmin on the ferroportin stability was impaired due to three distinct properties of the mutant ceruloplasmin: namely, a decreased ferroxidase activity, the mislocalization in the endoplasmic reticulum, and the failure of copper incorporation into apo-ceruloplasmin. Patients with aceruloplasminemia exhibited low serum hepcidin levels and a decreased ferroportin protein expression in the liver. The in vivo findings supported the notion that under low levels of hepcidin, mutant ceruloplasmin cannot stabilize ferroportin because of a loss-of-function in the ferroxidase activity, which has been reported to play an important role in the stability of ferroportin. The properties of mutant ceruloplasmin regarding the regulation of ferroportin may therefore provide a therapeutic strategy for aceruloplasminemia patients.

Prospective study of daily low-dose nedaplatin and continuous 5-fluorouracil infusion combined with radiation for the treatment of esophageal squamous cell carcinoma.

BACKGROUND: Protracted low-dose concurrent chemotherapy combined with radiation has been proposed for enhanced treatment results for esophageal cancer. We evaluated the efficacy and the toxicity of a novel regimen of daily low-dose nedaplatin (cis-diammine-glycolatoplatinum) and continuous infusion of 5-fluorouracil (5-FU) with radiation in patients with esophageal squamous cell carcinoma. METHODS: Between January 2003 and June 2008, 33 patients with clinical stage I to IVB esophageal squamous cell carcinoma were enrolled. Nedaplatin (10 mg/body/day) was administered daily and 5-FU (500 mg/body/day) was administered continuously for 20 days. Fractionated radiotherapy for a total dose of 50.4-66 Gy was administered together with chemotherapy. Additional chemotherapy with nedaplatin and 5-FU was optionally performed for a maximum of 5 courses after chemoradiotherapy. The primary end-point of this study was to evaluate the tumor response, and the secondary end-points were to evaluate the toxicity and the overall survival. RESULTS: Twenty-two patients (72.7%) completed the regimen of chemoradiotherapy. Twenty patients (60.6%) achieved a complete response, 10 patients (30.3%) a partial response. One patient (3.0%) had a stable disease, and 2 (6.1%) a progressive disease. The overall response rate was 90.9% (95% confidence interval: 75.7%-98.1%). For grade 3-4 toxicity, leukopenia was observed in 75.8% of the cases, thrombocytopenia in 24.2%, anemia in 9.1%, and esophagitis in 36.4%, while late grade 3-4 cardiac toxicity occurred in 6.1%. Additional chemotherapy was performed for 26 patients (78.8%) and the median number of courses was 3 (range, 1-5). The 1-, 2- and 3-year survival rates were 83.9%, 76.0% and 58.8%, respectively. The 1- and 2-year survival rates were 94.7% and 88.4% in patients with T1-3 M0 disease, and 66.2% and 55.2% in patients with T4/M1 disease. CONCLUSION: The treatment used in our study may yield a high complete response rate and better survival for each stage of esophageal squamous cell carcinoma. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT00197444.

Fecal cyclooxygenase 2 plus matrix metalloproteinase 7 mRNA assays as a marker for colorectal cancer screening.

We previously reported that fecal cyclooxygenase 2 (COX-2) mRNA assay, detecting COX-2 mRNA in feces, is useful for identifying subjects with colorectal cancer (CRC). To further improve the sensitivity, we evaluated the usefulness of the combination of COX-2 mRNA and matrix metalloproteinase 7 (MMP-7) mRNA assays as a marker of CRC. The study cohort included 62 patients with CRC and 29 control patients without colorectal neoplasia. RNA was isolated from routinely collected fecal samples. The expression levels of COX-2 and MMP-7 mRNAs were determined by nested reverse transcription-PCR. PCR conditions were optimized where the specificity of fecal COX-2 and MMP-7 mRNA assay result in 100%. The sensitivity of each fecal assay was 87% [95% confidence interval (95% CI), 76-94%] and 65% (95% CI, 51-76%) for CRC, respectively. The sensitivity of fecal RNA test (either marker being positive) was high for CRC (90%; 95% CI, 80-96%). The sensitivity of the fecal RNA test was also high (93%; 95% CI, 80-98%) in patients with stage I or II who are often cured by surgical resection. The fecal RNA test using COX-2 and MMP-7 mRNAs improved the sensitivity to detect CRC without decreasing the specificity. These results suggest that the fecal RNA test would be a promising approach for CRC screening, although larger clinical investigations are indicated.

A case of small bowel adenocarcinoma in a patient with Crohn's disease detected by PET/CT and double-balloon enteroscopy.

Small bowel adenocarcinoma (SBA) in patients with Crohn's disease (CD) is quite rare, difficult to diagnose without surgery, and has a poor prognosis. Here, we report a 48-year-old man with SBA and a 21-year history of CD who was diagnosed by a combination of positron emission tomography/computed tomography (PET/CT) and double-balloon enteroscopy (DBE). Since the age of 27 years, the patient had been treated for ileal CD and was referred to our hospital with persistent melena. Multiple hepatic tumors were found by CT. PET/CT detected an accumulation spot in the small bowel. DBE revealed an ulcerative tumor in the ileum about 100 cm from the ileocecal valve. An endoscopic forceps biopsy specimen showed poorly differentiated adenocarcinoma. There were some longitudinal ulcer scars near the tumor, and the chronic inflammation in the small bowel appeared to be associated with the cancer development. Previous reports suggest the risk of SBA in patients with CD is higher than in the overall population. Since early diagnosis is extremely difficult in these cases, novel techniques, such as PET/CT and DBE, may be expected to help in making a preoperative diagnosis of the development of SBA in CD.

Serum pepsinogens as an early diagnostic marker of H. pylori eradication.

BACKGROUND/AIMS: Helicobacter pylori (H. pylori) eradication increases the serum pepsinogen I/ pepsinogen II ratio and the percentage change in pepsinogen I/pepsinogen II ratios is a useful marker of H. pylori eradication. We studied whether the pepsinogen method could be an early diagnostic marker of H. pylori eradication even in patients persistently treated with a proton pump inhibitor. METHODOLOGY: Sixty-two H. pylori-positive patients underwent H. pylori-eradication therapy, followed by treatment with a proton pump inhibitor to cure ulcers. Serum levels of pepsinogen I and pepsinogen II were measured before, at the end of, and at 4 weeks after the eradication therapy. The cut-off values of percentage changes in pepsinogen I/pepsinogen II ratios for the diagnosis of eradication of H. pylori were set in proportion to pepsinogen I/pepsinogen II ratios before eradication in accordance with a previous report. RESULTS: Using the results of 13C-urea breath test as the standard, the sensitivity, specificity and validity of the pepsinogen method were 100.0%, 89.8% and 90.3%, respectively, at 4 weeks after eradication therapy. CONCLUSION: The percentage change in serum pepsinogen I/pepsinogen II ratios is useful as an early diagnostic marker for judgment of H. pylori eradication irrespective of proton pump inhibitor treatment.

Cyclooxygenase-2 and tumor biology.

There is now substantial evidence for the role of cyclooxygenase (COX)-2 in causation and prevention of cancer. Selective COX-2 inhibitors (coxibs) were considered attractive candidate chemoprevention agents; however, concerns over the toxicity of systemic selective inhibition have cast some doubt on COX-2 inhibition as a safe chemoprevention strategy. COX-2 can serve as a potential biomarker of tumor evaluation including prognosis. This chapter describes proposed mechanisms for the role of COX-2 in carcinogenesis, proliferation, inhibition of apoptosis, promotion of angiogenesis, enhanced invasiveness, immune modulation, and increased mutagenesis. Critical discussions focus on the use of COX-2 as a biomarker in the evaluation of neoplasm. Our chapter demonstrates that "Fecal COX-2 Assay," a novel method to detect COX-2 messenger RNA (mRNA) in feces from subjects with colorectal neoplasms, is potentially useful for colorectal cancer screening.

A deficiency of gastric interstitial cells of Cajal accompanied by decreased expression of neuronal nitric oxide synthase and substance P in patients with type 2 diabetes mellitus.

BACKGROUND: Gastrointestinal motility is impaired in patients with diabetes mellitus (DM). Interstitial cells of Cajal (ICC) in the gastrointestinal tract play a central role in gastrointestinal motility. The present study examined whether ICC density, or expression of neuronal nitric oxide synthase (nNOS)- and substance P (SP)-containing nerves in the gastric antrum, were altered in patients with type 2 DM. METHODS: Paraffin-embedded gastric specimens from 51 controls and 36 male DM patients with gastric cancer were used for immunohistochemistry. Serial sections were stained with Kit and mast cell tryptase-specific antibodies. Fresh-frozen gastric specimens from patients with gastric cancer were used for immunofluorescence. The specimens were stained with antibodies to Kit, nNOS, and SP, and levels of expression of these three markers were compared between controls and DM patients. RESULTS: ICC density in the inner circular muscle layer, but not in the myenteric plexus, was lower in patients with severe DM than in controls in paraffin-embedded specimens. In addition, decreased expression of nNOS and SP accompanied by reduced ICC density was observed in frozen specimens from patients with DM. CONCLUSIONS: These results suggest that lower gastric ICC, nNOS, and SP densities in patients with DM may be associated with the pathogenesis of diabetic gastroparesis.

Outcome of radiotherapy for gastric mucosa-associated lymphoid tissue lymphoma refractory to Helicobacter pylori eradication therapy.

BACKGROUNDS AND AIMS: Most low-grade gastric mucosa-associated lymphoid tissue (MALT) lymphomas respond to eradication of H. pylori, however, some are refractory. The effectiveness of radiotherapy for MALT lymphoma refractory to H. pylori eradication has been demonstrated. However, the long-term outcome and associated late radiation morbidity of radiotherapy for gastric MALT lymphoma are unclear. We evaluated the efficacy of radiotherapy and the radiation-associated adverse effects for gastric MALT lymphoma refractory to H. pylori eradication therapy. METHODS: Eighteen patients with H. pylori-positive gastric MALT lymphoma received H. pylori eradication therapy, of which three patients refractory to eradication of H. pylori were subsequently treated with irradiation (median 39 Gy, range 36-40 Gy). Efficacy of radiotherapy and associated radiation morbidity were evaluated. Adverse events were evaluated according to RTOG and NCI-CTC criteria. RESULTS: Radiation was well tolerated. All three patients treated with radiotherapy achieved complete remission, which was sustained for a median follow-up period of 42.0 months (range, 24-72 months). As acute radiation side effects, all patients experienced mild nausea (Grade 1 by RTOG and 1 point [1 category] in NCI-CTC). One patient had Grade 1 appetite loss. There were no severe late adverse events. One patient had left kidney atrophy of approximately 10%, but none had renal dysfunction. CONCLUSIONS: Radiotherapy for patients with gastric MALT lymphoma refractory to H pylori eradication was effective without any critical acute or late adverse events.