[Oxidative stress in porphyria and carriers].

UNLABELLED: We sought to establish a causal relationship between oxidative stress and porphyria in patients and carriers. We reported changes in urinary porphyrin concentrations related to 8-hydroxy-2'-deoxyguanosine. METHODS: We measured urinary 8-hydroxy-2'-deoxyguanosine concentration in porphyria patients and carriers with multifactorial inheritance as a possible marker of attack. The porphyria types included 10 patients with porphyria cutanea tarda, 5 with variegate porphyria, 8 with hereditary coproporphyria, 7 with congenital erythropoietic porphyria, 5 with erythropoietic protoporphyria, 5 with acute intermittent porphyria, 7 erythropoietic protoporphyria carriers, and 7 acute intermittent porphyria carriers. RESULTS: Urinary porphyrin concentrations in these patients were significantly higher than those in healthy subjects (p<0.001). Urinary 8-hydroxy-2'-deoxyguanosine concentrations were significantly high in dermatopathy porphyria types namely porphyria cutanea tarda (p<0.001), variegate porphyria (p<0.05), hereditary coproporphyria (p<0.05), congenital erythropoietic phyria (p<0.05), and erythropoietic protoporphyria (p<0.001). CONCLUSION: These results reveal that urinary 8-hydroxy-2'-deoxyguanosine concentration in cutis porphyria types is a good predictor of attack and abatement.

[A phase I study of postoperative combination therapy with TS-1 and low dose cisplatin against stage IV gastric carcinoma].

A dose-escalation study was conducted for postoperative patients with stage IV gastric cancer to determine the recommended dose of daily intravenous cisplatin combined with a fixed dose of TS-1. TS-1 was administered orally twice daily for 2 weeks followed by a 1-week rest. The dose of TS-1 was based on the body surface area (BSA) as follows: 80 mg/day for BSA less than 1.25, 100 mg/day for BSA 1.25 to less than 1.50, and 120 mg/day for BSA 1.5 or more. Three dose levels of cisplatin (2, 4, 6 mg/m(2)) were studied, and two courses were performed. Cisplatin was infused on day 1-5 and 8-12 for 30 minutes. The National Cancer Institute common toxicity criteria (NCI-CTC Version 3) were used to evaluate the grade of toxicity. Three patients enrolled in each level. Dose escalation was performed when dose-limiting toxicities (DLT) were seen in 0/3, and 3 more cases of the same level were added when DLTs were seen 1-2/3. Maximum-tolerated dose (MTD) were determined when DLTs were seen in 3 cases. DLTs were not recorded during the administration of CDDP up to 4 mg/m(2). However, DLTs were seen 3/3 at level 3. From these results, cisplatin of 4 mg/m(2)was determined to be the recommended dose (RD) in this protocol for postoperative stage IV gastric carcinoma.

Macrophage migration inhibitory factor promotes tumor invasion and metastasis via the Rho-dependent pathway.

PURPOSE: Macrophage migration inhibitory factor (MIF) plays an important role not only in the immune system but also in tumorigenesis. In this study, we investigated the potential role of MIF in association with tumor invasion and metastasis. METHODS: To assess the function of MIF, we knocked down the MIF mRNA using small interfering RNA (siRNA). Twenty-one base siRNA specific for the mRNA sequence of mouse MIF was introduced to a murine colon cancer cell line, colon 26. Tumor cell invasion was evaluated using a transwell method (8-microm pores) coated with Matrigel on the upperside membrane and with fibronectin on the underside membrane. Moreover, we investigated the signal transduction of lysophosphatidic acid (LPA) relevant to the Rho-dependent pathway and further examined the effect of MIF siRNA on this signal transduction system. In vivo, the tumor cells were pretreated with MIF siRNA and injected into the portal vein, and the effects on metastasis to the liver were evaluated. RESULTS: We found that MIF siRNA markedly reduced the invasion of the cells from the upperside to lowerside membranes. We revealed that the Rho-dependent pathway activated by LPA was suppressed by MIF siRNA. Next, we found that the tyrosine-phosphorylation of focal adhesion kinase and LPA-induced expressions of integrin beta1 were significantly suppressed by MIF siRNA. In vivo, metastasis to the liver was significantly inhibited by pretreatment of the cells with MIF siRNA. CONCLUSION: Taken together, these results suggest that MIF promotes tumor invasion and metastasis via the Rho-dependent pathway.

Evidence of dual function of macrophage migration inhibitory factor relevant to tumor progression and regression.

Macrophage migration inhibitory factor (MIF) is known to play an important role in broad-spectrum inflammation and immune responses. To evaluate the role of MIF in tumor growth, we established transgenic (Tg) mice (ICR strain) driven by cytomegalovirus (CMV) enhancer and beta-actin promoter. We inoculated Tg mice in the back with murine sarcoma cell line S-180 cells. The tumor growth rate was more enhanced in Tg mice than in littermate non-Tg mice up to day 9 after tumor inoculation. Surprisingly, most tumors embedded on the back of Tg mice regressed at day 10 after inoculation and eventually disappeared. Tumor volumes of non-Tg mice incessantly increased until death. We reinoculated the Tg mice with S-180 cells, which had been recovered from the first challenge, and found that the tumor cells were completely rejected in all cases. To identify the effector cells that eradicated the tumor cells, we prepared spleen cells from tumor-bearing Tg mice and carried out cell lysis assay. The magnitude of cytolytic activity of spleen cells obtained from Tg mice was significantly higher against S-180 cells, as well as natural killer cell-sensitive YAC-1 cells, than was the activity of cells from non-Tg mice. Furthermore, we observed that CTL activity of Tg mice against S-180 cells was significantly decreased by the deletion of CD8+ T cells or NK cells. On the other hand, the deletion of CD4+ cells minimally affected the cytolytic activity. Taken together, these results suggest that MIF has the potential to promote tumor growth and angiogenesis in the early phase and, by contrast, this protein could activate CD8+ cytotoxic T cells and NK cells, leading to tumor regression.

[Evaluation of low-dose FP therapy as adjuvant therapy for gastric cancer].

In the present study, we demonstrate the result of low-dose FP treatment as an adjuvant therapy for 30 patients of stage II or more progressive gastric cancer. 5-FU and CDDP were injected intravenously for 10 days from day 1 through day 5, and day 8 through 12 for 2 weeks at a dose of 250 mg/body and 10 mg/body, or for 14 days at a dose of 250 mg/m2/day and 5 mg/m2/day, respectively. Patients were excluded if they received less than 80% of the respective doses in a course of treatment by the protocol. This constituted a course of chemotherapy, which was repeated every 4 weeks. Grade 3 neutropenia was observed in one case. Other toxicities were anorexia, nausea, weight loss, diarrhea, general fatigue and elevation of serum creatinine, but they were not so severe. The two-year survival rate was 100% in cur A cases, 85% in cur B, and 0% in cur C. The median survival time of the cur C patients was 10 months. These results indicate that low-dose FP therapy is safe and recommendable for cur A and cur B patients. However, other treatment methods such as sequential chemotherapy are needed for cur C gastric cancer patients.

Porphyrias in Japan: compilation of all cases reported through 2002.

The first case of porphyria on record in Japan was a patient with congenital erythropoietic porphyria (CEP) reported by Sato and Takahashi in 1920. Since then until the end of December 2002, 827 cases of porphyrias have been diagnosed from characteristic clinical and/or laboratory findings (463 males, 358 females, and 6 of unknown sex). Essentially all inherited porphyrias have been found in Japan, with the incidences and clinical symptoms generally being similar to those reported for other countries. The male-female ratio was approximately 1:1 for CEP, whereas it was higher for erythropoietic protoporphyria. In contrast, preponderances of female patients exist with acute hepatic porphyrias, such as acute intermittent porphyria (AIP), variegate porphyria (VP), and hereditary coproporphyria (HCP), and with undefined acute porphyria. Although porphyria cutanea tarda (PCT) is believed to be increasing recently in women in other countries because of smoking and the use of contraceptives, it is still by far more prominent in males in Japan than in females. The recent increasing contribution of hepatitis C virus infection to PCT in Japan has also been recognized. but there have been no PCT cases in Japan with HFE gene mutations. Familial occurrence and consanguinity were high for CEP, as expected; however, significant consanguinity was also noted in families where CEP, AIP, HCP, VP, or PCT occurred as a single isolated case without a family history of disease. This survey also revealed that as many as 71% of acute hepatic porphyria cases were initially diagnosed as nonporphyria and later revised or corrected to porphyria, indicating the difficulty of diagnosing porphyria in the absence of specific laboratory testing for porphyrins and their precursors in urine, stool, plasma, and erythrocyte samples.

Induction of macrophage migration inhibitory factor by lysophosphatidic acid: relevance to tumor growth and angiogenesis.

Macrophage migration inhibitory factor (MIF) plays an important role not only in the immune system, but also in tumorigenesis. Lysophosphatidic acid (LPA), a unique lipid mediator, shares several biological functions with MIF, including promotion of tumor cell growth and associated angiogenesis. In this study, we investigated the signaling cross-talk between these two molecules during tumorigenesis and angiogenesis. We first examined the expression of MIF mRNA on a murine colon cancer cell line, colon 26, by LPA. We found that LPA enhanced the expression of MIF mRNA in a dose-dependent manner in vitro. In parallel, LPA stimulated cell growth and up-regulated the vascular endothelial growth factor (VEGF). These effects were dramatically blocked by 21 base double strand (ds) RNA specific for mouse MIF mRNA (RNAi). In vivo, colon 26 cells treated with MIF dsRNA were injected into the backs of mice. The size of tumor volumes became significantly smaller than that of controls. Angiogenesis examined by a Millipore chamber method was also suppressed by the MIF dsRNA. Next, we evaluated the signal transduction pathway relevant to the mitogen-activated protein kinase (MAPK) and Akt/PI3K pathways in response to LPA by RNAi. Ras activation and phosphorylation of Akt and ERK1/2 were strongly suppressed by the dsRNA. On the other hand, tyrosine phosphorylation was minimally changed by the treatment. Taken together, these results suggest that MIF could promote both tumor cell growth and angiogenesis induced by LPA via both the Ras-MAPK and Ras-Akt/PI3K signaling pathways.

[Keshan disease--a review from the aspect of history and etiology].

OBJECTIVE: Keshan disease is an endemic cardiomyopathy found in Keshan, north-east China. The first patient was identified in 1935. This disease is characterized by a blood circulation disorder, endocardium abnormality and myocardium necrosis. Selenium (Se) deficiency is thought to be a major factor by Chinese scientists. However, the exact etiology has not been clarified up to now. The government decided to apply sodium selenite to growing crops, and the incidence of the disease decreased dramatically. However, a few cases still occur as chronic or latent types. This paper reviews Keshan disease from a historical aspect and also the present situation. METHODS: We made a reference survey and summarised the etiology, pathological changes, clinical manifestation, and other views of Keshan disease. RESULT: So far, epidemiological surveys have shown that Keshan disease occurs in large areas in 14 provinces in China, mainly in the countryside. It has been confirmed by clinical and pathological studies that Keshan disease is an independent endemic myocardial disease caused by biological and geological factors. The largest prevalence age rates are boys under 15 years old and women of childbearing age. There are several hypotheses; acute carbon monoxide poisoning, virus infection, malnutrition, or selenium deficiency. The first is not currently believed to be the cause. The following was pointed out; studies on the relationship between diet and the endemic areas of Keshan disease in 1961, where the food custom of the local residents was relatively simple and a so-called "one-sided diet" (eating a limited variety of food) might be related to the incidence of Keshan disease. In 1973, large-scale investigations on the natural environments were performed in the endemic areas of Keshan disease in the whole country. As a result, it was reported that there was a relationship between the incidence of Keshan disease and the special natural environment in the endemic areas and the cause of Keshan disease was strongly supported by nutritional, biological, geological and chemical (selenium deficiency) factors. In 1981, on the other hand, it was found that the levels of antibodies against Coxsackie virus were higher in the serum of Keshan disease patients than of normal subjects. This fact supposed that the cause of Keshan disease was related to a virus infection. However, it is difficult to explain why the clinical and pathological manifestations of Keshan disease are similar to those of other diseases, e.g. idiopathic dilatational myocardial disease. Further research should be performed on Keshan disease to clarify the etiology.

[A family of acute intermittent porphyria].

A 62-year-old man who had twice received laparotomies for abdominal pain of unknown origin was admitted to our hospital with acute abdominal pain. His family history of acute intermittent porphyria (AIP) suggested that it arose from acute porphyria. We treated the patient with 5% glucose solution by i.v. drip infusion and his abdominal pain improved rapidly. Diagnosis of AIP was established by the demonstration of reduced erythrocyte porphobilinogen deaminase (PBGD) activity and a point mutation (CAG --> CGG) in a splicing site in intron 10/exon 11 in the PBGD gene by DNA analysis. For screening of AIP carriers in his family, we measured erythrocyte PBGD activity. Four of his seven children were successfully diagnosed as AIP carriers. This is the ninth AIP family report, in which a mutation in the PBGD gene was revealed by DNA analysis.

[Neoadjuvant chemoradiotherapy for advanced squamous cell carcinoma of thoracic esophagus].

Advanced thoracic esophageal cancer has a poor prognosis despite advances in surgery, such as three-field lymph node dissection. Multimodal therapy is needed to improve local control, resectability and survival rate. Fifteen patients with advanced squamous cell carcinoma of the thoracic esophagus were treated with neoadjuvant chemoradiotherapy (NAC) combined with concurrent radiation (30 Gy/12 f) and 3 courses of 5-FU and CDDP (CDDP 5 mg/m2/day + 5-FU 250 mg/m2/day: day 1-5: div). In the absence of unresectable disease and surgical risk, 12 patients underwent esophagectomy (Group 1) and 3 patients underwent additional chemoradiotherapy because of high surgical risk (Group 2). Side effects consisted of nausea, vomiting and myelo-suppression in 8 patients, but all patients tolerated and completed a full course of NAC. The effective rate (CR + PR) of NAC was 58.3% in Group 1 and 66.7% in Group 2. No patients showed pathological CR. Two-year survival rate was 28.1% in Group 1 (PR: 33.3%, NC: 20.0%) and 33.5% in Group 2. This protocol had acceptable toxicities but did not show survival benefit. Further trials are necessary to improve survival rate.

[Modified pharmacokinetic modulation chemotherapy (PMC) with medication of UFT and intraarterial infusion of 5-FU for advanced unresectable HCC].

UNLABELLED: Advanced unresectable hepatocellular carcinoma (HCC) was treated with modified pharmacokinetic modulation chemotherapy (PMC). METHOD: Modified PMC consists of medication with UFT and intraarterial infusion of 5-FU. The dose of UFT is 300 or 400 mg/day. The infusion to hepatic artery of 5-FU is performed with 500 mg/body in an outpatient clinic once a week from reservoir port for 5 hours. RESULTS: The number of recurrent cases after hepatectomy was 5, and that of initial cases with unresectable HCC was 3. Three cases had tumor thrombus in the main portal branch. One patient had tumor thrombus in the inferior vena cava, which reached to the right atrium. The mean number of infusions in all cases was 21. One case showed PR, and 3 cases NC. Three of 6 mortality cases died from liver failure without tumor progression. One year survival rates of the patients with tumor thrombus in the portal trunk or IVC were 75.0%. The mean survival period of these cases was 12.5 +/- 4.2 months. CONCLUSION: Modified PMC had no severe side effect and was effective for advanced unresectable HCC.

Exogenous coproporphyrin III production by Corynebacterium aurimucosum and Microbacterium oxydans in erythrasma lesions.

Erythrasma is a superficial skin disease caused by Gram-positive Corynebacterium species. Coral-red fluorescence under Wood's light, strongly suggestive of erythrasma, can be attributed to the presence of porphyrins. Fractionated porphyrin analysis in erythrasma lesions is yet to be reported. We attempted to investigate erythrasma lesions by isolating the responsible bacteria and determining their exogenous porphyrin production by HPLC analysis. We observed a 78-year-old woman with erythrasma who had a well-demarcated slightly scaling patch on her left foot, between the fourth and fifth toes. Two kinds of colonies on 5 % sheep blood agar were obtained from this lesion. Analysis of the 16S rRNA sequence revealed the colonies to be Corynebacterium aurimucosum and Microbacterium oxydans. HPLC analysis demonstrated that coproporphyrin III (Copro III) levels were clearly elevated, although the amounts of protoporphyrin were diminished. These results indicate that the fluorescent substance was Copro III. This study supports the view that excess Copro III synthesis by C. aurimucosum and M. oxydans leads to accumulation of porphyrin in cutaneous tissue, which emits a coral-red fluorescence when exposed to Wood's light.

Array comparative genomic hybridization analysis revealed four genomic prognostic biomarkers for primary gastric cancers.

Unlike the case with some other solid tumors, whole genome array screening has not revealed prognostic genetic aberrations in primary gastric cancer. Comparative genomic hybridization (CGH) using bacterial artificial chromosome (BAC) arrays for 56 primary gastric cancers resulted in identification of four prognostic loci in this study: 6q21 (harboring FOXO3A; previously FKHRL1), 9q32 (UGCG), 17q21.1 approximately q21.2 (CASC3), and 17q21.32 (HOXB3 through HOXB9). If any one of these four loci was deleted, the prognosis of the patient was significantly worse (P = 0.0019). This was true even for advanced tumors (stage IIIA, IIB, or IV, n = 39) (P = 0.0113), whereas only 1 of the 17 patients with less advanced tumors (stage IA, IB, or II; n = 17) died of disease after surgery. Multivariate analysis according to the status of four BACs or pathological stage based on the Japanese Classification of Gastric Carcinoma (stages IA, IB, and II vs. stages IIIA, IIIB, and IV) demonstrated that the BAC clone status was also an independent prognostic factor (P = 0.006). These findings may help predict which patients with malignant potential need more intensive therapy, or may point to new therapeutic approaches especially for advanced tumors. The parameter here termed the integrated genomic prognostic biomarker may therefore be of clinical utility as a prognostic biomarker.

Adenine nucleotide translocator transports haem precursors into mitochondria.

Haem is a prosthetic group for haem proteins, which play an essential role in oxygen transport, respiration, signal transduction, and detoxification. In haem biosynthesis, the haem precursor protoporphyrin IX (PP IX) must be accumulated into the mitochondrial matrix across the inner membrane, but its mechanism is largely unclear. Here we show that adenine nucleotide translocator (ANT), the inner membrane transporter, contributes to haem biosynthesis by facilitating mitochondrial accumulation of its precursors. We identified that haem and PP IX specifically bind to ANT. Mitochondrial uptake of PP IX was inhibited by ADP, a known substrate of ANT. Conversely, ADP uptake into mitochondria was competitively inhibited by haem and its precursors, suggesting that haem-related porphyrins are accumulated into mitochondria via ANT. Furthermore, disruption of the ANT genes in yeast resulted in a reduction of haem biosynthesis by blocking the translocation of haem precursors into the matrix. Our results represent a new model that ANT plays a crucial role in haem biosynthesis by facilitating accumulation of its precursors into the mitochondrial matrix.