Effects of micro-encapsulation on morphology and endocrine function of cryopreserved neonatal porcine islet-like cell clusters.

BACKGROUND: For the success of clinical islets transplantation, the development of a long-term storage method is necessary. However, the structure of digested islets is scanty for culture and cryopreservation. In this study, the effect of micro-encapsulation to cryopreserved porcine islet-like cell clusters (ICCs) was investigated. METHODS: The ICCs prepared from neonatal pigs by collagenase digestion and culture technique were cryopreserved and micro-encapsulated in 5% agarose membranes. After cryopreservation, ICC cultured without encapsulation (group A) and cultured with encapsulation (group B) were assessed by comparison with no cryopreserved ICC (control) both in vitro by static incubation test and in vivo in a xenotransplantation study. RESULTS: Micro-encapsulation was able to maintain the fine morphology and the number of ICCs of group B after 7 days of culture. There were not significant differences in insulin secretion of group B and control on day 1 and 7 of culture (1 day:11+/-0.99, 7 days: 5.30+/-1.08 microU/ICC/hr NS versus control). On day 7 of culture, the retrieval rate of group B (105.2+/-9.8%) is obviously higher compared with group A (63.0+/-6.3%). In the xenotransplatation model, the ICCs of group B showed long survival time (7.9+/-0.4 weeks) and good transplantation effect. CONCLUSION: Our study suggests that micro-encapsulation is one of the useful method for cryopreserved ICC to maintain the fine morphology and effectively recover the endocrine function.

Efficacy of continuous hyperthermic peritoneal perfusion for the prophylaxis and treatment of peritoneal metastasis of advanced gastric cancer: evaluation by multivariate regression analysis.

Thirty-two patients with advanced gastric cancer underwent continuous hyperthermic peritoneal perfusion (CHPP) combined with surgery: to prevent peritoneal recurrence in 15 patients without peritoneal metastasis (prophylactic CHPP) and to treat 17 patients with peritoneal metastases (therapeutic CHPP). The postoperative outcome was compared with that of control patients treated with surgery alone. Peritoneal recurrence was less frequent (26%) and the 5-year survival rate was significantly higher (39%) in the patients with prophylactic CHPP than in 40 control patients (42 and 17%, respectively). The patients with therapeutic CHPP showed significantly better median survival than did 20 control patients (11 vs. 6 months). Cox multivariate regression analysis revealed that CHPP was an independent prognostic factor in the prophylactic study (hazard ratio = 0.3965), and that the independent prognostic factor in the therapeutic study was not CHPP but complete resection of the peritoneal metastasis. Thus, CHPP has no marked benefit for established peritoneal metastasis. CHPP for the prevention of peritoneal recurrence may have a beneficial effect on long-term survival, but a prospective randomized trial is needed to clarify its prognostic value.

Surgical results of pancreaticoduodenectomy for carcinoma of the distal third of the stomach.

Pancreaticoduodenectomy (PD) was performed as a radical operation on 10 patients who had stage III or IV carcinoma of the distal third of the stomach which invaded the pancreas head (T4) or had level 3 lymph node metastasis. The surgical results of the PD were compared with those of 69 patients treated with subtotal gastrectomy (SG). Although the postoperative morbidity was higher (70%) in the PD group than in the SG group (32%), no hospital death occurred. The overall postoperative survival provided by PD was as good as that provided by SG for 43 patients who had stage III or IV tumors (the 5-year survival rates, 40 versus 45%). Regarding the T4 tumors invading the pancreas, the survival of the 9 patients with PD was better than that of the 12 patients with SG (median survival time, 19 versus 9 months). Thus, PD might improve the postoperative survival of patients with carcinoma of the distal stomach invading the pancreas.

Expression of DCC protein in colorectal tumors and its relationship to tumor progression and metastasis.

A loss of heterozygosity (LOH) at the DCC gene locus was detected in colorectal tumors, and this LOH might be related to metastasis. The aim of this study was to determine DCC protein expression in colorectal cancer and to evaluate its prognostic value. Allelic loss of the DCC locus was observed in 16 of the 23 patients (66.7%). In all 16 patients with LOH, DCC expression was decreased in the cancer tissue compared with the adjacent normal mucosa. All 23 colorectal tumors had decreased expression of this protein relative to the adjacent normal colonic mucosa in Western blot analysis. The levels of DCC protein were significantly lower in cancer tissues than in adenoma tissues. Decreased DCC protein expression was also observed by immunohistochemistry in the colorectal cancer cases. There were significant correlations between DCC protein expression and histologic type, venous invasion, and hematogenous metastasis. Patients with DCC-protein-negative tumors had a greater relative risk of recurrence compared with those whose tumors were DCC protein-positive. The 5-year survival rate was 91.0% in patients with DCC-protein-positive tumors, and 58.8% in those with DCC-protein-negative tumors; these differences between the two groups of patients were significant (p < 0.01). In multivariate analysis using the Cox regression model, DCC protein expression emerged as an independent prognostic indicator. These findings suggested that a decrease in DCC expression may have an important role in the progression of colorectal cancers and may be a biologic marker of prognostic significance.

[A case of stage IV (N4) esophageal cancer successfully treated with concurrent chemoradiotherapy].

A 71-year-old man with inoperable intrathoracic esophageal cancer was treated by concurrent chemoradiation. A dose of 48 Gy (neck) and 60.6 Gy (mediastinum) and four courses of 5-FU (500 mg/day)-CDDP (50 mg/week) were delivered. The esophageal tumor and metastatic lymph nodes of the neck showed a complete response (CR) to the treatment, whereas paraaortic lymph nodes evidenced no change (NC). The patient is doing well without symptoms at 13 months after treatment. The chemoradiotherapy produced effective improvement and quality of life in this patient.

Clinical significance of combined immunohistochemical detection of CD44v and sialyl LeX expression for colorectal cancer patients undergoing curative resection.

To evaluate their prognostic value, the expressions of CD44v and sialyl LeX (SLX) in colorectal cancers were studied immunohistochemically. Tissue specimens were reacted with monoclonal antibodies (mAb) CD44-1V and CSLEX-1. Of the 145 colorectal cancer patients undergoing curative resection, 59 (40.7%) were positive for mAb CD44-1V, and 40 (27.6%) were positive for mAb CSLEX-1. There was a significant correlation between the combined expression of SLX and CD44v8-10 and lymph node metastasis. The patients with tumors negative for CD44v8-10 and SLX had the most favorable prognoses. Conversely, the patients with tumors positive for both CD44v8-10 and SLX had a high recurrence rate and the poorest prognoses. In a multivariate analysis using the Cox regression model, the combined expression of SLX and CD44v8-10 emerged as an independent prognostic indicator. These results suggested that the combined expression of CD44v8-10 and SLX may be a biologic marker of prognostic significance.

Clinical significance of serum levels of CD44 variant exons 8-10 protein in colorectal cancer.

We examined serum levels of a CD44 splice variant that contained variant exons 8-10 (CD44v8-10) as a tumor marker in colorectal cancer patients. We performed enzyme-linked immunosorbent assays in 81 sera obtained from 71 colorectal cancer patients and 10 healthy controls. Serum CD44v8-10 levels were significantly higher in the colorectal cancer patients than in the healthy controls (0.209 +/- 0.098 versus 0.114 +/- 0.019 OD; P < 0.01). There was a close correlation between immunohistochemical expression and serum CD44v8-10 levels. Surgical resection of the tumors resulted in a reduction of serum CD44v8-10 levels. There was no significant correlation between serum CD44v8-10 level and serosal invasion or histologic type. However, a significant correlation was observed between serum CD44v8-10 level and lymphatic or venous invasion. In addition, serum CD44v8-10 levels were significantly higher in carcinomas associated with lymph node or liver metastasis than in those without metastasis. These findings suggest the usefulness of serum CD44v8-10 level in the prediction of colorectal cancer metastasis.

Prognostic value of DNA ploidy and proliferating cell nuclear antigen in gastric cancer.

DNA ploidy and the labeling index (LI) of proliferating cell nuclear antigen (PCNA) in gastric cancers were determined using cytofluorometry and immunohistochemistry, respectively, and the prognostic value of these two parameters was evaluated. Of 66 patients with advanced gastric cancers treated with radical resection, 27 with aneuploidy and a high PCNA-LI (> or = 40) showed the lowest 5-year survival rate (38%). Twenty patients with diploid cancers and a high LI showed a lower 5-year survival rate (59%) than the 15 patients with diploid cancers and a low LI (<40), who had the highest 5-year survival rate (86%). A multivariate analysis showed that the grouping based on the ploidy and the LI was an independent prognostic factor. Thus, the combination of DNA ploidy and PCNA-LI may be a useful prognostic indicator for advanced gastric cancers.

Expression of nm23-H1 gene and Sialyl Lewis X antigen in breast cancer.

The nm23 gene has been proposed as a candidate tumor metastasis suppressor in some human cancers. Sialyl Lewis X (sLex) has been demonstrated to play an important role in the adhesion of human cancer cells to human vascular endothelium, inducing metastasis. Little information has been reported about the correlation between the expression of nm23 and sialylated carbohydrate antigens. In the present study, 102 surgically resected primary breast cancer tissues were sectioned and stained with antibody against nm23-H1 and sLex. Of the 102 cases, 39 (38.2%) cases with a reduced expression of nm23-H1 were observed, and the numbers of sLex-positive cases were 61 (59.8%), respectively. The reduced expression of nm23-H1 and the positive expression of sLex were significantly associated with lymph node involvement. Among the 100 patients who underwent curative surgery, the disease-free survival rate was significantly correlated to both the nm23-H1 and sLex expressions. No interrelated expressions were found between nm23-H1 and sLex. In multivariate analysis using Cox regression model, combination assay of nm23-H1 and sLex expression emerged as independent significant prognostic factors. These results suggest that nm23-H 1 gene and sLex may be involved in different steps of the metastatic process in human breast cancer, and immunohistochemical detection of the combination of sLex and nm23-H1 may be a biologic marker of prognostic significance.

[Telomerase activity in colorectal carcinoma and its relationship to clinicopathological findings].

Telomerase is thought to be responsible for cell immortality. The telomerase activity in carcinomas has been remarked since 1995. We examined telomerase activity in colorectal carcinoma by TRAP (Telomeric repeat amplification protocol) assay, and investigated its relationship to clinicopathological findings. We could analyse telomerase activities in 33 cases (66%) of 50 colorectal carcinomas, whereas the activity was not found in all 13 cases of noncancerous colorectal mucosa. There was no relation between the telomerase activity and the clinicopathological findings or metastatic status. We confirmed telomerase activities in much of colorectal carcinomas in spite of their progression. The carcinoma cells might be immortal from their early stage of progression by means of telomerase activity.

Usefulness of metyrapone treatment to suppress cancer metastasis facilitated by surgical stress.

BACKGROUND: One causative factor of tumor metastasis enhanced by surgical stress is thought to be hypersecretion of endogenous glucocorticoids. This study evaluated the effectiveness of metyrapone treatment and adrenalectomy in preventing the harmful effects of glucocorticoids in the enhancement of tumor metastasis resulting from surgical stress. METHODS: The effect of dexamethasone on pulmonary metastasis of MRMT-1 cells and on the number of peripheral lymphocytes was evaluated in rats. To evaluate the suppressive effect of adrenalectomy and metyrapone treatment on operation-induced enhancement of metastasis, several parameters such as induction of pulmonary metastasis, serum corticosterone levels, and the number of blood lymphocytes and apoptotic thymocytes were determined. RESULTS: With dexamethasone treatment, the number of peripheral lymphocytes rapidly decreased; in contrast, pulmonary metastasis increased. The serum corticosterone level was doubled at 1 hour, apoptotic thymocyte numbers were increased about sevenfold at 3 hours and about fourfold at 6 hours, and blood lymphocyte numbers were decreased at 3 hours after laparotomy, which facilitated about a 10-fold increase in the pulmonary metastasis. These changes were almost completely suppressed by preoperative adrenalectomy and metyrapone treatment. CONCLUSIONS: Preoperative metyrapone treatment, which suppresses hypersecretion of endogenous glucocorticoids as a result of operation, modulates the enhancement of cancer metastases and may be an effective treatment.

Preservation of pancreatic islets in rodent models.

Islet-cell transplantation has some advantages over vascularized pancreas transplantation, but data from clinical islet cells transplantation have shown that some serious problems need to be overcome. One of them is the storage of islet cells. We investigated methods of preserving islet cells using culture-preservation and cryopreservation. Cryopreservation is thought to be effective for long-term preservation of large quantities of islet cells, because they can be cryopreserved without loss of their physiologic activity using a relatively rapid cooling rate of 25 degrees C/min. Moreover, in the allogenic transplantation models of cryopreservation of dissociated islet cells, there was a significant prolongation of survival time. These results suggest that cryopreservation of islet cells involves not only variable preservation methods but may also lead to a modification of graft immunogenicity.

Reduced expression of deleted colorectal carcinoma (DCC) protein in established colon cancers.

Using a bacterial fusion protein, a deleted colorectal carcinoma (DCC)-specific monoclonal antibody (MAb) 127-22 was established. Although MAb 127-22 reacted with almost all normal tissues, it did not react or only weakly reacted with many cancer cell lines, including colonic cancer lines, in flow cytometry. In Western immunoblots, the MAb reacted with a single 190-kDa molecule in a myeloma line Ara-10 extract. This component was scarcely detected in colonic cancer cell lines. Immunoblots of samples from 25 pairs of colonic cancers and adjacent normal tissues and from five adenoma tissues revealed that all normal colonic and adenoma tissues significantly expressed the DCC protein, whereas colonic cancer tissues showed poor expression. These results indicate not only deletion of and lowered mRNA expression of the DCC gene, but also marked reduction of DCC protein occurred in colonic cancer tissues. In addition, colonic cancer patients with liver metastasis expressed significantly lower levels of DCC than those without, suggesting the prognostic value of DCC expression.

The role of lymph node dissection in the treatment of gallbladder carcinoma.

BACKGROUND: Lymph node involvement is an important prognostic factor in gallbladder carcinoma (GBC). The lymph node involvement pattern, extent, and indications for systematic lymph node dissection for patients with advanced GBC were investigated. METHODS: Forty-one patients with GBC who underwent radical resection with systematic regional lymph node dissection over the past 11 years were analyzed. RESULTS: The lymph node metastasis rate was 63.4% overall, 0% in pT1 disease, 61.9% in pT2 disease, and 81.3% in pT3/pT4 disease. When reviewed according to site, the rate was 41.5% in pericholedochal lymph nodes, 22.0% in the lymph nodes around the common hepatic artery and the portal vein, 36.6% in the posterior pancreaticoduodenal lymph nodes, 28% (5/18) in the celiac lymph nodes, 19% (3/ 16) in the superior mesenteric artery (SMA) lymph nodes, and 26% (7/27) in the aortocaval paraaortic lymph nodes. Patients with severe hepatoduodenal ligament invasion had high rates of paraaortic lymph node involvement. The mortality rate was 2.4% (1 of 41 patients) and the 5-year survival rate was 33.1% overall, 100% in patients with pT1 disease, 49.8% in patients with pT2 disease, and 0% in patients with pT3/pT4 disease. The 5-year survival rate for pT2 disease according to lymph node involvement was 72.7% in patients with pN0+ pN1+ positive posterior pancreaticoduodenal lymph nodes and positive lymph nodes around the common hepatic artery in the N2 patients and 0% in the patients with positive celiac and SMA lymph nodes in the N2 patient group or the positive paraaortic lymph node group (P < 0.05). CONCLUSIONS: These results suggest that systemic dissection of N1 lymph nodes, posterior pancreaticoduodenal lymph nodes, and lymph nodes around the common hepatic artery and the portal vein in N2 patients is necessary to improve the prognosis of those patients with pT2 disease without moderate or severe hepatoduodenal ligament invasion.

Inhibition of liver metastasis formation by anti-CD44 variant exon 9 monoclonal antibody.

The overexpression of variants of the glycoprotein CD44 is thought to be associated with the tumorigenesis and progression of human cancers. We examined the role of the variant CD44v8-10 in the metastasis of the human colon cancer cell line HT29 using a monoclonal antibody (mAb 44-1V) reactive with the v9 product. After immunization with mAb 44-1V, the growth of HT29m cells in vitro was not retarded. Six-to 8-week-old mice were divided into 4 groups for liver metastasis assay. All animals in control groups injected with intrasplenic HT29m developed metastases. In contrast, only one of the animals injected with HT29m that reacted with mAb 44-1V developed a metastatic tumor in the liver. The intravenous administration of mAb 44-1V after intrasplenic HT29m injection did not inhibit the formation of liver metastasis. In addition, the adhesiveness of the HT29m cells to the basement membrane matrix was decreased by treatment with the anti-CD44v9 mAb. These findings indicated that a CD44 variant containing the products of variants of exons v8-10 may play an important role in adhesion of tumor cells to the capillaries of distant organs in the metastatic process.

Bcl-2 protein expression in breast cancer and its relationship to prognosis.

The expression of bcl-2 protein was studied in invasive breast cancer by immunohistochemistry. Fourty-six (56.8%) bcl-2 protein-positive tumors were found in 81 breast cancers. There was no significant correlation between the bcl-2 protein immunoreactivity and histologic type, primary tumor status, or lymph node metastasis. However, a strong positive relationship was demonstrated between bcl-2 immunoreactivity and estrogen receptor status. The 5-year survival rate and disease-free survival rates were 73.7% and 71.1% of patients with bcl-2-positive tumors, and 62.5% and 58.0% of those with bcl-2-negative tumors; these differences between the two groups of patients wete significant (p<0.05). In multivariate analysis using Cox regression model, bcl-2 immunoreactivity emerged as an independent prognostic indicator in breast cancer patients.

Lectin-histochemical reactivity of sialic acid in breast cancer and its relationship to prognosis using limulus polyphemus agglutinin.

Studies of circulating sialic acid have revealed its relationship with a variety of malignant tumors. It is not vet clear whether sialic acid could be used as a prognostic marker of breast cancer, and few studies have examined sialic acid expression in the cell membrane and cytoplasm of breast cancer cells by means of the lectin-histochemical technique. In the present study, we used biotinylated limulus polyphemus agglutinin (LPA), a special binding lectin of sialic acid, to stain sialic acid in breast cancer cells. Of the 104 cases of breast cancer examined, 59 (56.7%) positive cases were observed. There was a significant correlation between the LPA staining and the clinicopathologic features of all patients, including pathological stage and lymph node metastasis. Among the 100 patients who underwent curative operation, the mean disease-free survival rate of the 45 patients who were LPA-negative was significantly higher than that of the 55 LPA-positive patients (p<0.05). These results suggest that the positive expression of sialic acid in breast cancer could be used as a marker of malignancy potential, as well as a poor survival factor, and the biotinylated LPA assay may provide a convenient and useful method to predict the prognosis of breast cancer.

Significance of tumor marker SLX in colorectal cancer.

We examined serum SLX for its significance as a tumor marker in 109 colorectal cancer patients. There a close correlation with immunohistochemical expression of SLX and serum SLX level. Serum SLX was positive in 16.5% of 109 patients with colorectal cancers. There was no significant correlation between serum SLX level and histologic type or primary tumor status. There were significant correlations between serum SLX positive rates and both lymph node and hematogenous metastasis. In 7 SLX positive cases who underwent curative resection, 4 patients had already recurrence in the liver. Our findings suggest that serum SLX values may be a biologic marker of metastasis.