Berberine Induces Apoptotic Cell Death via Activation of Caspase-3 and -8 in HL-60 Human Leukemia Cells: Nuclear Localization and Structure-Activity Relationships.

Berberine (BBR), an isoquinoline alkaloid, is a well-known bioactive compound contained in medicinal plants used in traditional and folk medicines. In this study, we investigated the subcellular localization and the apoptotic mechanisms of BBR were elucidated. First, we confirmed the incorporation of BBR into the cell visually. BBR showed antiproliferative activity and promptly localized to the nucleus from 5[Formula: see text]min to 15[Formula: see text]min after BBR treatment in HL-60 human promyelocytic leukemia cells. Next, we examined the antiproliferative activity of BBR (1) and its biosynthetically related compounds (2-7) in HL-60 cells. BBR exerted strongest antiproliferative activity among 1-7 and the results of structures and activity relation suggested that a methylenedioxyl group in ring A, an [Formula: see text]-alkyl group at C-9 position, and the frame of isoquinoline may be necessary for antiproliferative activity. Moreover, BBR showed the most potent antiproliferative activity in HL-60 cells among human cancer and normal cell lines tested. Next, we examined the effect of BBR on molecular events known as apoptosis induction. In HL-60 cells, BBR induced chromatin condensation and DNA fragmentation, and triggered the activation of PARP, caspase-3 and caspase-8 without the activation of caspase-9. BBR-induced DNA fragmentation was abolished by pretreatment with inhibitors against caspase-3 and caspase-8, but not against caspase-9. ERK and p38 were promptly phosphorylated after 15 min of BBR treatment, and this was correlated with time of localization to the nucleus of BBR. These results demonstrated that BBR translocated into nucleus immediately after treatments and induced apoptotic cell death by activation of caspase-3 and caspase-8.

Correlation between meropenem and doripenem use density and the incidence of carbapenem-resistant Pseudomonas aeruginosa.

Optimal use of carbapenems is an important issue in the prevention of resistance in Pseudomonas aeruginosa. In this study, we investigated the correlation between antimicrobial use density (AUD) of carbapenems and imipenem/cilastatin (IPM/CS) or meropenem (MEPM) susceptibility of P. aeruginosa strains. The AUD of five carbapenems [IPM/CS, panipenem/betamipron, biapenem, MEPM and doripenem (DRPM)] was examined every 6 months between 2006 and 2008. The AUD was calculated using the defined daily doses methodology developed by the World Health Organisation. A minimum inhibitory concentration of IPM/CS or MEPM of < or =4 mg/L was considered to be sensitive. There was a significant negative correlation between MEPM susceptibility and the total AUD of MEPM and DRPM [r=-0.823, 95% confidence interval (CI) -0.035 to -0.980; P=0.044]. Furthermore, there was a significant correlation between MEPM susceptibility and IPM/CS susceptibility (r=0.839, 95% CI 0.084 to 0.981; P=0.037). Cross-resistance was therefore investigated and only 5.6% of MEPM-insensitive strains were susceptible to IPM/CS, although 43.3% of IPM/CS-insensitive strains were susceptible to MEPM. These results suggest that curtailing the use of MEPM and DRPM may curb the emergence not only of MEPM-resistant strains but also IPM/CS-resistant strains.

Effect of methotrexate treatment on expression levels of organic anion transporter polypeptide 2, P-glycoprotein and bile salt export pump in rats.

High-dose methotrexate (HDMTX) chemotherapy with leucovorin (LV) rescue has been used as a therapeutic strategy in oncology since the 1970s. Adverse reactions following extension of methotrexate (MTX) elimination are a crucial problem in HDMTX chemotherapy. MTX is a substrate for drug transporters, which are multidrug resistance protein 2 (Mrp2), organic anion transporter polypeptide 2 (Oatp2) and other transporters. We previously reported that MTX treatment downregulated the expression level of Mrp2 in rats. Here we examined the effect of MTX treatment on the expression of Oatp2, P-glycoprotein (P-gp) and bile salt export pump (Bsep) in rats. MTX was single-injected intraperitoneally at a dose of 150 mg/kg, and Western blot analysis was performed. The levels of Oatp2, P-gp and Bsep in the liver on day 4 after treatment were downregulated to 36.3 +/- 6.9%, 51.5 +/- 5.2% and 61.8 +/- 5.5% (mean +/- S.E.M.) of controls, respectively. Expression levels of P-gp in the kidney and ileum were also downregulated to 38.5 +/- 1.6% and 16.2 +/- 1.6% of controls, respectively. These effects of MTX were partially recovered by LV, which rescues normal cells from MTX toxicity. In conclusion, the result indicates that MTX treatment downregulates expression levels of Oatp2, P-gp and Bsep.

Effect of pre-treatment with St John's Wort on nephrotoxicity of cisplatin in rats.

An herbal health care supplement, St John's Wort (SJW, Hypericum perforatum) has become widely used in the treatment of depression, and is known to interact with therapeutic drugs. Here we report a preventive effect of SJW on cisplatin nephrotoxicity in rats. Rats were given SJW (400 mg/kg/day, p.o.) for 10 consecutive days, and were injected with cisplatin (5 mg/kg, i.v.) on the day after the final SJW treatment. Cisplatin treatment increased the serum creatinine level, which is an index of nephrotoxicity, to 1.51+/-0.22 mg/dl (mean+/-SE) from 0.28+/-0.05 mg/dl (control) on day 5 after the cisplatin injection. This increase fell significantly to 0.86+/-0.13 mg/dl by pre-treatment with SJW. Cisplatin-induced histological abnormality of the kidney was blocked by pre-treatment with SJW. When SJW was administered for 10 days, the amounts of renal metallothionein (MT) and hepatic multidrug resistance protein 2 (Mrp2) were increased to 164.8+/-13.0% and 220.8+/-39.3% (mean+/-SE) of controls, respectively. GSH levels in the kidney and liver were not changed. Total and free cisplatin concentration in serum was not influenced by SJW treatment. In conclusion, the results suggest that pre-treatment with SJW may diminish cisplatin nephrotoxicity.

Influence of preoperative administration of omega-3 fatty acid-enriched supplement on inflammatory and immune responses in patients undergoing major surgery for cancer.

OBJECTIVE: Polyunsaturated fatty acid supplementation may produce beneficial effects after surgery. We investigated the influence of preoperative administration of a supplement rich in arginine, omega-3 fatty acids, and RNA, Impact (Japan), on inflammatory and immune responses in patients undergoing major surgery for cancer. METHODS: Patients in the supplement group (n = 12) received 1 L/d of Impact (Japan) for 5 d before surgery, and those in the control group (n = 14) received an ordinary diet without Impact (Japan) before surgery. Plasma levels of omega-3 and omega-6 fatty acids, thromboxane B(2), prostaglandin E(2), inflammatory markers, nutritional markers, cytokines, and cytokine receptors were obtained 5 d before the operation at the starting point of supplementation in the supplement group. Samples were collected on postoperative days (PODs) 0, 1, 3, and 7. RESULTS: After taking the supplement, significant increases in omega-3 fatty acids and rapid turnover proteins were found the day after ending supplementation (POD-0), whereas thromboxane B(2) levels and the ratio of omega-6 fatty acids to omega-3 fatty acids were significantly lower than before supplementation (P < 0.001). On POD-0 only, inflammatory markers and cytokine receptors in the supplement group showed low levels in comparison with the control group (P < 0.05). On POD-1 and POD-3, remarkable decreases in polymorphonuclear leukocyte-elastase and interleukin-8 in the supplement group were observed. CONCLUSION: Our findings suggest that oral administration of a supplement rich in omega-3 fatty acids for 5 d before surgery may improve not only preoperative nutritional status but also preoperative and postoperative inflammatory and immune responses in patients who have cancer.

HTLV-1 provirus load in peripheral blood lymphocytes of HTLV-1 carriers is diminished by green tea drinking.

Human T-cell lymphotropic virus type 1 (HTLV-1) is causatively associated with adult T-cell leukemia (ATL) and HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). Since a high level of HTLV-1 provirus load in circulating lymphocytes is thought to be a risk for ATL and HAM/TSP, diminution of HTLV-1 provirus load in the circulation may prevent these intractable diseases. Our previous study (Jpn J Cancer Res 2000; 91: 34-40) demonstrated that green tea polyphenols inhibit in vitro growth of ATL cells, as well as HTLV-1-infected T-cells. The present study aimed to investigate the in vivo effect of green tea polyphenols on HTLV-1 provirus load in peripheral blood lymphocytes on HTLV-1 carriers. We recruited 83 asymptomatic HTLV-1 carriers to examine HTLV-1 provirus DNA with or without administration of capsulated green tea extract powder. Thirty-seven subjects were followed up for 5 months by measuring HTLV-1 provirus load after daily intake of 9 capsules of green tea extract powder per day (equivalent to 10 cups of regular green tea), and 46 subjects lived ad libitum without intake of any green tea capsule. The real-time PCR quantification of HTLV-1 DNA revealed a wide range of variation of HTLV-1 provirus load among asymptomatic HTLV-1 carriers (0.2-200.2 copies of HTLV-1 provirus load per 1000 peripheral blood lymphocytes). Daily intake of the capsulated green tea for 5 months significantly diminished the HTLV-1 provirus load as compared with the controls (P = 0.031). These results suggest that green tea drinking suppresses proliferation of HTLV-1-infected lymphocytes in vivo.