The efficacy of conditioned medium released by tonsil-derived mesenchymal stem cells in a chronic murine colitis model.

Tonsil-derived mesenchymal stem cells (TMSC) have characteristics of MSC and have many advantages. In our previous studies, intraperitoneal (IP) injection of TMSC in acute and chronic colitis mouse models improved the disease activity index, colon length, and the expression levels of proinflammatory cytokines. However, TMSC were not observed to migrate to the inflammation site in the intestine. The aim of this study was to verify the therapeutic effect of conditioned medium (CM) released by TMSC (TMSC-CM) in a mouse model of dextran sulfate sodium (DSS)-induced chronic colitis. TMSC-CM was used after seeding 5×105 cells onto a 100 mm dish and culturing for 5-7 days. TMSC-CM was concentrated (TMSC-CM-conc) by three times using a 100 kDa cut-off centrifugal filter. Seven-week-old C57BL/6 mice were randomly assigned to the following 5 groups: 1) normal, 2) colitis, 3) TMSC, 4) TMSC-CM, and 5) TMSC-CM-conc. Chronic colitis was induced by continuous oral administration of 1.5% dextran sulfate sodium (DSS) for 5 days, followed by 5 additional days of tap water feeding. This cycle was repeated two more times (total 30 days). Phosphate buffered saline (in the colitis group), TMSC, TMSC-CM, and TMSC-CM-conc were injected via IP route 4, 4, 12, and 4 times, respectively. Reduction of disease activity index, weight gain, recovery of colon length, and decreased in the expression level of the proinflammatory cytokines, interleukin (IL)-1ß, IL-6, and IL-17 were observed at day 30 in the treatment groups, compared to control. However, histological colitis scoring and the expression level of tumor necrosis factor α and IL-10 did not differ significantly between each group. TMSC-CM showed an equivalent effect to TMSC related to the improvement of inflammation in the chronic colitis mouse model. The data obtained support the use of TMSC-CM to treat inflammatory bowel disease without any cell transplantation.

Emergence of rifampin-resistant staphylococci after rifaximin administration in cirrhotic patients.

OBJECTIVES: Rifaximin, a poorly absorbed antibiotics, has gut-specific therapeutic effects. Although frequently prescribed to manipulate intestinal luminal bacterial population in various diseases, the possible induction of antibacterial cross-resistance to a target pathogen is a major concern in long-term rifaximin administration. We aimed to evaluate whether rifampin-resistant staphylococci could evolve after rifaximin treatment in cirrhotic patients. METHOD: A total of 25 cirrhotic patients who were administered rifaximin for the prevention of hepatic encephalopathy were enrolled. Swabs from both hands and the perianal skin were acquired on day 0 (before rifaximin treatment), period 1 (1-7 weeks after treatment), and period 2 (8-16 weeks after treatment) the staphylococcal strain identification and rifampin-resistance testing. RESULTS: A total of 198 staphylococcal isolates from 15 species were identified. Staphylococcus epidermidis was isolated most frequently, and Staphylococcus haemolyticus was the most common resistant species both from hands and perianal skin. Eleven patients (44.0%) developed rifampin-resistant staphylococcal isolates in period 1. Among these patients, only six (54.5%) were found to have rifampin-resistant isolates in period 2, with no significant infectious events. Rifampin-resistant staphylococcal isolates were more frequently found in perianal skin than from the hands. No patients acquired a newly resistant strain in period 2. CONCLUSIONS: About one-half of cirrhotic patients in this study developed rifampin-resistant staphylococcal isolates after rifaximin treatment. Although the resistant strains were no longer detected in about half of the patients in the short-term, the long-term influence of this drug treatment should be determined.

Therapeutic potential of tonsil-derived mesenchymal stem cells in dextran sulfate sodium-induced experimental murine colitis.

The therapeutic potential of tonsil-derived mesenchymal stem cells (TMSC) prepared from human tonsillar tissue has been studied in animal models for several diseases such as hepatic injury, hypoparathyroidism, diabetes and muscle dystrophy. In this study, we examined the therapeutic effects of TMSC in a dextran sulfate sodium (DSS)-induced colitis model. TMSC were injected in DSS-induced colitis mice via intraperitoneal injection twice (TMSC[x2]) or four times (TMSC[x4]). Control mice were injected with either phosphate-buffered saline or human embryonic kidney 293 cells. Body weight, stool condition and disease activity index (DAI) were examined daily. Colon length, histologic grading, and mRNA expression of pro-inflammatory cytokines, interleukin 1ß (IL-1ß), IL-6, IL-17 and tumor necrosis factor α, and anti-inflammatory cytokines, IL-10, IL-11 and IL-13, were also measured. Our results showed a significant improvement in survival rates and body weight gain in colitis mice injected with TMSC[x2] or TMSC[x4]. Injection with TMSC also significantly decreased DAI scores throughout the experimental period; at the end of experiment, almost complete reversal of DAI scores to normal was found in colitis mice treated with TMSC[x4]. Colon length was also significantly recovered in colitis mice treated with TMSC[x4]. However, histopathological alterations induced by DSS treatment were not apparently improved by injection with TMSC. Finally, treatment with TMSC[x4] significantly reversed the mRNA levels of IL-1ß and IL-6, although expression of all pro-inflammatory cytokines tested was induced in colitis mice. Under our experimental conditions, however, no apparent alterations in the mRNA levels of all the anti-inflammatory cytokines tested were found. In conclusion, our findings demonstrate that multiple injections with TMSC produced a therapeutic effect in a mouse model of DSS-induced colitis.

Significance of calcium-sensing receptor expression in gastric cancer.

OBJECTIVE: The calcium-sensing receptor (CaSR) is known to have differential expression in various carcinomas and normal tissues. It has been shown to be involved in carcinogenesis or tumor suppression. However, its role in gastric cancer remains unknown. This study was performed to determine the CaSR expression level in gastric cancer and non-tumor gastric tissues and to examine the related clinicopathological factors. MATERIALS AND METHODS: Thirty-one pairs of gastric cancer tissues and matched non-tumor gastric tissues were obtained from surgical tissues after gastrectomy. Using real-time polymerase chain reaction, we measured CaSR mRNA expression. We evaluated the association between CaSR mRNA expression and clinicopathological variables based on the downregulation or upregulation of CaSR mRNA expression in gastric cancer tissues compared to those of matched non-tumor gastric tissues. By immunohistochemistry, we confirmed CaSR expression levels in gastric cancer tissues. RESULTS: Downregulation of CaSR mRNA was observed in 77.4% of gastric cancer tissues compared to their matched normal tissues. Downregulated CaSR was associated with a tendency for deeper invasion into the proper muscle (p = 0.028) and more advanced stage (II-IV; p = 0.012). CONCLUSION: We conclude that downregulation of CaSR may contribute to the prevention or suppression of tumor outgrowth.

Involvement of adiponectin in early stage of colorectal carcinogenesis.

BACKGROUND: Although altered levels of adiponectin have been reported as a potential risk factor in colorectal cancer (CRC), the importance of the role played by adiponectin in colorectal carcinogenesis has not been established. We sought to examine the expression pattern of adiponectin and adiponectin receptors (AdipoRs) in the normal-adenoma-carcinoma sequence and to assess the implications of adiponectin in colorectal carcinogenesis. METHODS: Serum adiponectin concentrations, and the mRNA and protein expression of adiponectin and AdipoRs were examined using serum and tissues from patients with CRC, advanced adenoma, and a normal colon. mRNA expression of AdipoRs and epithelial-mesenchymal transition regulators including E-cadherin, cyclooxygenase-2 (COX-2) and T-cadherin were examined in HCT116 cells treated with adiponectin. RESULTS: Serum adiponectin concentrations in patients with advanced adenoma and CRC were lower than those in controls. Adiponectin mRNA was not detected in colonic tissue, whereas AdipoRs mRNA was lower in advanced adenoma and CRC than that in normal colon tissues. Immunohistochemical staining demonstrated that adiponectin was expressed in spindle-shaped cells of the subepithelial layer in normal colon tissues, whereas ill-defined overexpression of adiponectin was seen in the stroma of advanced adenoma and CRC tissues. AdipoRs expression was strong in normal epithelium, but weak to negative in the epithelia of CRC tissues. Adiponectin downregulated COX-2 mRNA expression in vitro, but upregulated T-cadherin in HCT116 cells. CONCLUSIONS: Systemic adiponectin and local AdipoRs expression in the colon may be associated with anti-tumorigenesis during the early stages of CRC. These findings offer new insight into understanding the relationship between adiponectin and colorectal carcinogenesis.

Significance of preoperative tissue levels of vascular-endothelial cadherin, liver-intestine cadherin and vascular endothelial growth factor in gastric cancer.

BACKGROUND/AIMS: The aims of this study were to examine the expressions of endothelium specific VE-cadherin, intestine specific LI-cadherin, and vascular endothelial growth factor (VEGF), and to determine their relationships with the clinicopathological parameters of gastric cancer. METHODS: A total 47 patients with gastric cancer who underwent surgery were enrolled. Endoscopic biopsies were obtained from the cancer and normal mucosa, respectively. Using semiquantitative RT-PCR, the mRNA expression levels of VE-cadherin, LI-cadherin and VEGF were measured by tumor/normal (T/N) ratios. The protein expressions of VE-cadherin, LI-cadherin and VEGF were examined by Western blot and immunohistochemical stain in surgically resected tissues. The clinicopathological variables were reviewed and analyzed, retrospectively. RESULTS: Twenty two cases (46.8%) of VE-cadherin, 25 cases (53.2%) of LI-cadherin and 27 cases (51.1%) of VEGF mRNA expressions were overexpressed in gastric cancer compared to normal tissue. There was a tendency for T/N ratio of VE-cadherin mRNA to correlate with the lymphatic invasion (p=0.07) and the lymph node metastasis (p=0.099) in advanced gastric cancer. The T/N ratio of LI-cadherin mRNA showed significant association with distant metastasis (p=0.031) and lymphatic invasion especially in advanced gastric cancer (p=0.023). There was a tendency for the T/N ratio of VEGF mRNA to correlate with the distant metastasis (p=0.073) in advanced gastric cancer. CONCLUSIONS: As increased mRNA expression of LI-cadherin was associated with distant metastasis and lymphatic invasion especially in the biopsy specimen of advanced gastric cancer before surgery, it may provide useful preoperative information on tumor aggressiveness.

Methylation of the Tumor Suppressor Gene RUNX3 in Human Gastric Carcinoma.

BACKGROUND/AIMS: RUNX3 (PEBP2alphaC/CBFA3/AML2) is a novel tumor suppressor gene in the human gastric carcinoma. The aims of this study were to determine the methylation of RUNX3 promoter and the association between RUNX3 methylation and the clinicopathological characteristics of patients with gastric carcinoma. METHODS: Seventy-nine patients with gastric carcinoma were studied prospectively from April 2005 to May 2007. The methylations of RUNX3 promoter on the gastric carcinoma specimens and the corresponding nonneoplastic mucosa were evaluated by methylation-specific polymerase chain reaction. RESULTS: Comparison of the results with the clinicopathological characteristics identified RUNX3 monoallelic methylation in 32.9% (26/79) of the gastric carcinoma patients and in 11.4% (9/79) of those with nonneoplastic mucosa (p=0.053). The monoallelic methylated gastric carcinoma specimens predominantly consisted of well- and moderately differentiated carcinomas (44.7%), with the unmethylated group constituting 22.0% of them (p=0.031). Among the 48 patients (60.8%) who underwent gastrectomy, there was no correlation between the two groups with regard to Lauren's classification (p=0.235), depth of invasion (p=0.990), nodal status (p=0.601), stage (p=0.900), lymphatic invasion (p=0.537), and vascular invasion (p=0.815). CONCLUSIONS: Methylation of the tumor suppressor gene RUNX3 might be one of the mechanisms involved in the pathogenesis of gastric carcinoma.

Clinical significance of tissue levels of matrix metalloproteinases and tissue inhibitors of metalloproteinases in gastric cancer.

BACKGROUND: Matrix metalloproteinases (MMPs) are one of the major classes of proteolytic enzymes involved in tumor invasion and metastasis, being inhibited by naturally occurring tissue inhibitors of metalloproteinases (TIMPs). In this study, we examined the expression of MMP-2, MMP-9, membrane-type 1 (MT1)-MMP, TIMP-1, and TIMP-2 in biopsy tissues of gastric cancer, and the correlation between their expression and clinicopathological parameters. METHODS: Biopsy specimens from 66 patients with gastric carcinoma were available for this study. To determine the expression of MMP-2, MMP-9, MT1-MMP, TIMP-1, and TIMP-2, semiquantitative reverse transcriptase-polymerase chain reaction (RT-PCR) was carried out on tumor and normal tissues, respectively, sampled during diagnostic gastroscopic examination. Immunohistochemical staining of representative samples using monoclonal antibody directed against MT1-MMP was done, and the clinicopathological variables were reviewed retrospectively. RESULTS: The expression level of MMPs and TIMPs was evaluated using the tumor : normal (T/N) ratios of MMPs and TIMPs. The T/N ratio of MT1-MMP mRNA showed a significant correlation with lymph node metastasis and tumor stage (P < 0.05). The other RT-PCR data of MMP-2, MMP-9, TIMP-1, and TIMP-2 did not show any significant correlation with clinicopathological parameters. Immunohistochemistry for MT1-MMP showed a positive immunoreaction in gastric adenocarcinoma and negative staining in normal mucosa. CONCLUSIONS: The correlation between the increased expression of MT1-MMP and clinicopathological variables reflects a role in predicting the aggressive behavior of gastric cancer. Because an RT-PCR assay can be performed on biopsy specimens obtained before surgery, an evaluation of MT1-MMP expression in biopsy specimens by RT-PCR may provide useful preoperative information on tumor aggressiveness.

[Expression of MMP-2, HIF-1alpha and VEGF in colon adenoma and colon cancer].

BACKGROUND/AIMS: This study was aimed to investigate the expression of matrix metalloproteinase-2 (MMP-2), hypoxia-inducible factor (HIF)-1alpha, and vascular endothelial growth factor (VEGF) in colonic adenoma-carcinoma sequence. METHODS: Thirty-two tissue samples of colon adenoma, 11 of early colon cancer and 36 of advanced colon cancer were collected by colonoscopic biopsy. Normal colonic tissues were also collected from the same subjects. The mRNA expression levels of MMP-2, HIF-1alpha, and VEGF were quantitated using semiquantitative reverse transcription polymerase chain reactions. The protein expressions of activated MMP-2 and HIF-1alpha were examined by gelatin zymography and by Western blot in surgically resected cases, respectively. RESULTS: The expression level of MMP-2 mRNA showed a progressive increase in the advance of the colorectal adenoma-carcinoma sequence (p<0.05). In colon cancer tissues, the expression level of MMP-2 mRNA showed an increasing trend according to differentiation, lymphatic invasion and Dukes' stage (p<0.05). The protein expression of activated MMP-2 was observed in 10 of 11 (91%) cases of cancer tissues. The mRNA expression levels of HIF-1alpha and VEGF were greater in tissues of early and advanced colon cancer compared with colon adenoma (p<0.05; p<0.001). The protein expression of HIF-1alpha was observed in 9 of 11 (82%) cases of cancer tissues. The mRNA expression level of HIF-1alpha showed a positive correlation with MMP-2 and VEGF, respectively (r=0.52, p<0.001; r=0.76, p<0.001). CONCLUSIONS: MMP-2, HIF-1alpha, and VEGF may be useful in detecting early carcinogenesis and progression of colon cancer.

Overview of administrative simplification provisions of HIPAA.

The administrative simplification provisions of the Health Insurance Portability and Accountability Act (HIPAA) of 1996 are considered to have a considerable impact on the health care industry in the United States of America. The HIPAA-mandated electronic transactions using Electronic Data Interchange (henceforth EDI) methods have brought significant issues on privacy, confidentiality and security for individually identifiable information not to mention technical issues. The purpose of this paper is to facilitate the understanding of those issues surrounding the administrative simplification part of HIPAA. Toward this goal, the paper provides an overview of HIPAA through a systematic approach of understanding its evolution and its requirements.