Tumoral presence of human cytomegalovirus is associated with shorter disease-free survival in elderly patients with colorectal cancer and higher levels of intratumoral interleukin-17.

Infectious diseases are closely related to cancer. Human cytomegalovirus (HCMV) has been implicated in the promotion of tumour growth, and is present in the tumour specimens of colorectal cancer (CRC). This study aimed to investigate whether tumoral presence of HCMV is associated with a different clinical outcome in elderly patients with CRC. We analysed archived tumour specimens from 95 CRC patients aged ≥65 years. HCMV was detected by PCR. Clinical, pathological, disease-free and overall survival data were compared between patients with HCMV-positive and HCMV-negative tumours. A quantitative RT-PCR array was used to evaluate the expression levels of cytokines genes of T-helper subpopulations in tumours. In the Kaplan-Meier analysis of the 81 patients who underwent curative surgery, 39 patients with HCMV-positive tumours had a lower disease-free survival rate (p 0.024). For patients with stage II or stage III tumours, tumoral HCMV status correlated with disease-free survival more closely than the traditional histopathological staging methods. In a multivariate Cox proportional hazard model, tumoral presence of HCMV independently predicted tumour recurrence in 5 years (hazard ratio 4.42; 95% CI 1.54-12.69, p 0.006). The qRT-PCR analysis of ten stage II tumours showed that the gene expression levels of interleukin-17-the signature cytokine of T-helper 17 cells-and its receptor, interleukin-17 receptor C, were higher in the five HCMV-positive tumours. Our results suggest that the presence of HCMV in CRC is associated with poorer outcome in elderly patients. How the virus interacts with the tumour microenvironment should be further investigated.

Patients with nonalcoholic fatty liver disease have higher risk of colorectal adenoma after negative baseline colonoscopy.

AIM: The study aimed to determine whether nonalcoholic fatty liver disease (NAFLD) is an independent risk factor of adenoma after negative baseline colonoscopy. METHOD: A retrospective cohort study was conducted on 1522 health-check individuals who underwent two consecutive colonoscopies at Taipei Veterans General Hospital between 2003 and 2010. Those developing an adenoma after an initial negative baseline colonoscopy (adenoma group) were compared with those in whom the second colonoscopy was negative (nonadenoma group). Anthropometric measurements, biochemical tests and the presence of NAFLD were compared between the two groups. RESULTS: The adenoma group had a higher prevalence of NAFLD than the nonadenoma group (55.6% vs 38.8%; P < 0.05). On multivariate logistic regression analysis, NAFLD was an independent risk factor (OR = 1.45, 95% CI: 1.07-1.98) for adenoma formation after a negative baseline colonoscopy. The risk of colorectal adenoma increased when NAFLD patients had other morbidities including metabolic syndrome, hypertension or smoking (OR = 2.85, 4.03 and 4.17). CONCLUSION: NAFLD is an independent risk factor for colorectal adenoma formation after a negative baseline colonoscopy. The risk is higher in individuals with NAFLD and other comorbidities, such as hypertension, smoking or metabolic syndrome.

Gastroesophageal and laryngopharyngeal reflux profiles in patients with obstructive sleep apnea/hypopnea syndrome as determined by combined multichannel intraluminal impedance-pH monitoring.

BACKGROUND: The profiles of gastroesophageal reflux (GER) and laryngopharyngeal reflux (LPR) in patients with obstructive sleep apnea/hypopnea syndrome (OSAHS) have never been explored. The aim of the study was to investigate the reflux profile in OSAHS patients. METHODS: Consecutive snoring out-patients suspected with having OSAHS and 20 healthy volunteers were included. All subjects underwent simultaneous 24-h combined multichannel intraluminal impedance-pH (MII-pH) monitoring and polysomnography. Obstructive sleep apnea/hypopnea syndrome was defined when the apnea/hypopnea index was over 5. Stepwise multiple logistic regression analysis was performed to determine the predictor for OSAHS. KEY RESULTS: Fifty-three patients were included, 37 with and 16 without OSAHS. The prevalence of reflux symptoms was similar between OSAHS (35.1%) and non-OSHAS (37.5%) patients. More OSAHS patients, compared with non-OSAHS patients and healthy volunteers, had pathologic acid GER, nocturnal acid GER, and prolonged acid clearance (P < 0.001). However, no difference in non-acid reflux episodes was observed among the three groups. Laryngopharyngeal reflux was detected in 51.4%, 43.8%, and 35.0% of OSAHS, non-OSAHS, and healthy volunteers, respectively (P = 0.034). In OSAHS patients, there was no difference in the sleep parameters between patients with and without LPR. Body mass index was the only predictor of OSAHS in the regression analysis. CONCLUSIONS & INFERENCES: OSAHS patients have more pathologic acid GER and prolonged acid clearance than non-OSAHS patients whereas non-acid reflux was similar between the two groups. However, BMI, not GER, is the only independent predictor for OSAHS. Laryngopharyngeal reflux occurs in more than half of OSAHS patients despite no significant association with OSAHS.

Predictors of acid suppression success in patients with chronic laryngitis.

BACKGROUND: Up to 50% of the patients suspected of reflux laryngitis syndrome failed to respond to acid suppression therapy. However, predictors of acid suppression success have not been determined. METHODS: Consecutive patients with chronic laryngitis were enrolled prospectively. All the patients underwent laryngoscopy, esophagogastroduodenoscopy and 24-h multichannel intraluminal impedance and pH (MII-pH) monitoring before receiving rabeprazole 10 mg b.i.d. for 3 months. Patient was considered as a responder to acid suppression if the chief laryngeal complaint score during the last week since last interview had decreased by at least 50% after the start of therapy compared with baseline. Cox regression analysis was used to determine the independent predictors of acid suppression success. KEY RESULTS: Of 92 patients (age 42.4 ± 14.3 years, 50 women), 42 (45.7%) responded to acid suppression after 3 months. Gastroesophageal reflux disease was defined in 22 patients, of whom 19 patients had pathological distal esophageal acid exposure and 5 were defined as erosive esophagitis. The time to response showed a significant hazard ratio for patients with increased distal esophageal acid exposure time (ß: 0.93; HR: 2.55; 95% CI: 1.24-5.24; P = 0.011) and increased laryngopharyngeal bolus exposure time (BET; ß: 0.96; HR: 2.61; 95% CI: 1.36-5.00; P = 0.004). The latter had the best Youden Index (0.34) and accuracy (68.5%). CONCLUSIONS & INFERENCES: The success of acid suppression on chronic laryngitis could be predicted using reflux parameters detected by MII-pH, among which increased laryngopharyngeal BET is the best.

Interleukin-1 receptor associated kinases-1/4 inhibition protects against acute hypoxia/ischemia-induced neuronal injury in vivo and in vitro.

Neuronal Toll-like receptors (TLRs)-2 and -4 have been shown to play a pivotal role in ischemic brain injury, and the interleukin-1 receptor associated kinases (IRAKs) are considered to be the key signaling molecules involved downstream of TLRs. Here, we investigated the expression levels of IRAK-1 and -4 and the effects of IRAK-1/4 inhibition on brain ischemic insult and neuronal hypoxia-induced injury. Male Sprague-Dawley (SD) rats and the rat neuroblastoma B35 cell line were used in these experiments. Permanent middle cerebral artery occlusion (MCAO) was induced by the intraluminal filament technique, and B35 cells were stimulated with the hypoxia-mimetic, cobalt chloride (CoCl(2)). Following induction of hypoxia/ischemia (H/I), B35 cells and cerebral cortical neurons expressed higher levels of IRAK-1 and -4. Furthermore, IRAK-1/4 inhibition decreased the mortality rate, functional deficits, and ischemic infarct volume by 7 days after MCAO. Similarly, IRAK-1/4 inhibition attenuated CoCl(2)-induced cytotoxicity and apoptosis in B35 cells in vitro. Our results show that IRAK-1/4 inhibition decreased the nuclear translocation of the nuclear factor-kappaB (NF-κB) p65 subunit, the levels of activated (phosphorylated) c-jun N-terminal kinase (JNK) and cleaved caspase-3, and the secretion of TNF-α and IL-6 in B35 cells at 6 h after CoCl(2) treatment. These data suggest that IRAK-1/4 inhibition plays a neuroprotective role in H/I-induced brain injury.

VEGF -460T → C polymorphism and its association with VEGF expression and outcome to FOLFOX-4 treatment in patients with colorectal carcinoma.

The -460T → C polymorphism of vascular endothelial growth factor (VEGF) gene significantly increases its promoter activity. A pilot study was conducted to assess the influence of this polymorphism on clinicopathological features of patients with colorectal carcinoma. In total, 228 patients were enrolled, including 100 with stage II/III colorectal carcinoma receiving curative surgery and 128 with metastatic disease. An excellent correlation in VEGF -460 genotypes based on white blood cells and tumor tissues existed, but there was no between-group difference in patients with or without colorectal carcinoma. A marked increase in intratumor and circulating VEGF levels were observed in patients with the T/C or C/C genotypes (P < 0.01), which was associated with increased extent of invasion, nodal involvement, poor histological differentiation, subsequent metastasis and shorter survival in stage II/III patients treated with curative surgery (P < 0.01). For patients with metastatic disease, this polymorphism was associated with a lower response rate to FOLFOX-4 (P = 0.03) and shorter survival (P < 0.001). By multivariate analysis, this polymorphism was identified as an independent prognostic factor (P = 0.01). These data suggest that -460T → C polymorphism of VEGF gene, by increasing VEGF expression and subsequent angiogenesis, could be a key determinant for increased tumor recurrence and a poor prognosis of patients with colorectal carcinoma. However, this study is exploratory and is not adjusted for multiple comparisons, requiring independent replication.

Primary vs. delayed resection for obstructive left-sided colorectal cancer: impact of surgery on patient outcome.

PURPOSE: By comparing surgical outcomes between primary and delayed resection, we addressed whether and how surgical strategies impacted prognosis of patients with left-sided colorectal cancer underwent emergency curative resection. METHODS: Between January 1980 and December 2002, a total of 143 patients were identified who presented with obstructive left-sided colorectal cancer and received emergency curative resection in Taipei Veterans General Hospital. Patients were stratified according to the timing of tumor resection into two groups: primary resection and delayed resection. Demographic data of the patients, characteristics of the tumors, and short-term and long-term outcomes were analyzed and compared between the two groups. RESULTS: The demographic data and tumor characteristics did not differ between the two groups except for more rectal cancers in the delayed resection group (P=0.021). Primary resection group had a higher anastomotic leakage rate (P=0.017) and a trend toward a higher mortality rate, which did not reach statistical significance (P=0.063). The median follow-up intervals were similar (60.4 vs. 58.3 months; P=0.79). The median survival tended to be longer in delayed resection group (66 vs. 105 months; P=0.088). Overall five-year and ten-year survival for primary resection were 43.7 and 31.9 percent, respectively, compared with 67.2 and 53.2 percent, respectively, for delayed resection. CONCLUSIONS: Delayed resection seems to be a safer procedure and provided a better oncologic outcome compared with primary resection in obstructive left-sided colorectal cancer under emergency situations.

System to assess accuracy of micromanipulation.

The authors had previously developed an optical micro motion sensing system (M2S2) using a pair of orthogonally placed position sensitive detectors (PSD) to track 3D displacement of the tip of a microsurgical instrument in real-time. In the M2S2 system, an infrared (IR) diode is used to illuminate the workspace. A ball is attached to the tip of an intraocular shaft to reflect IR rays onto the PSDs. Instrument tip position is then calculated from the centroid position of reflected IR light on the PSDs. The M2S2 system together with a test platform is used as an evaluation system to assess the accuracy and physiological tremor of subjects performing micromanipulation tasks. Since the need to use the ball at the instrument tip prevents the subjects from performing the manipulation tasks precisely, a laser light is provided as a guide for the subjects to aim at the target precisely. A laser diode module is placed inside the instrument to provide the required laser light. The instrument intraocular shaft is replaced with a same-sized hollow tube to let the laser light from the instrument pass through down to the target. The laser light spot position on the platform is used to access the performance of the subjects. The laser light spot position is calculated from the tilt angle information provided by an accelerometer placed inside the instrument, and the instrument tip position information given by M2S2.

Increasing live donor kidney transplantation: a randomized controlled trial of a home-based educational intervention.

With the shortage of deceased donor kidneys and the superior clinical outcomes possible with live donor kidney transplantation (LDKT), more patients should seriously consider LDKT. However, little is known about how best to educate patients and their family members about LDKT. We evaluated the effectiveness of a home-based (HB) educational program in increasing LDKT. Patients were randomized to clinic-based (CB) education alone (CB, n = 69) or CB plus HB education (CB+HB, n = 63). Compared to CB, more patients in the CB+HB group had living donor inquiries (63.8% vs. 82.5%, p = 0.019) and evaluations (34.8% vs. 60.3%, p = 0.005) and LDKTs (30.4% vs. 52.4%, p = 0.013). Assignment to the CB+HB group, White race, more LDKT knowledge, higher willingness to discuss LDKT with others, and fewer LDKT concerns were predictors of having LDKT (p-values < 0.05). Both groups demonstrated an increase in LDKT knowledge after the CB education, but CB+HB led to an additional increase in LDKT knowledge (p < 0.0001) and in willingness to discuss LDKT with others (p < 0.0001), and a decrease in LDKT concerns (p < 0.0001). Results indicate that an HB outreach program is more effective in increasing LDKT rates than CB education alone.

Suppression of fatty acid synthase in MCF-7 breast cancer cells by tea and tea polyphenols: a possible mechanism for their hypolipidemic effects.

Tea is a heavily consumed beverage world wide because of its unique aroma, less cost and broad availability. Fatty acid synthase (FAS) is a key enzyme in lipogenesis. FAS is overexpressed in the malignant human breast carcinoma MCF-7 cells and its expression is further enhanced by the epidermal growth factor (EGF). The EGF-induced expression of FAS was inhibited by green and black tea extracts. The expression of FAS was also suppressed by the tea polyphenol (-)-epigallocatechin 3-gallate (EGCG), theaflavin (TF-1), TF-2 and theaflavin 3,3'-digallate(TF-3) at both protein and mRNA levels that may lead to the inhibition of cell lipogenesis and proliferation. Both EGCG and TF-3 inhibit the activation of Akt and block the binding of Sp-1 to its target site. Furthermore, the EGF-induced biosyntheses of lipids and cell proliferation were significantly suppressed by EGCG and TF-3. These findings suggest that tea polyphenols suppress FAS expression by downregulating EGF receptor/PI3K/Akt/Sp-1 signal transduction pathway, and tea and tea polyphenols might induce hypolipidemic and antiproliferative effects by suppressing FAS.

Gasless laparoscopy-assisted colorectal surgery.

BACKGROUND: Laparoscopy has gained wide acceptance as a treatment modality in a variety of colonic and rectal disorders. Currently, most laparoscopic procedures are performed using a carbon dioxide (CO2) pneumoperitoneum, which can lead to cardiopulmonary loading and subsequent complications. The object of this study was to assess the feasibility of gasless laparoscopy-assisted colorectal surgery (GLACS) as an alternative method. METHODS: Patients with benign colonic lesions were enrolled in the study. The operative field was exposed with a subcutaneous wire lifting system. A small incision, ~5 cm in length, was made early in the operation. The surgeon operated through the trocar ports and this incision using both laparoscopic and conventional instruments. The cardiopulmonary responses of the patients were monitored continuously during the operation. RESULTS: Fifteen consecutive patients underwent GLACS. In two patients (13.3%), conversion to open surgery was necessary. The exposure and ease of the procedure were acceptable. However, when the patients were stratified into hemicolectomy and sigmoidectomy groups, GLACS scored more favorably in the sigmoidectomy group. There were no operative deaths. One minor complication developed postoperatively. All of the patients recovered uneventfully, with return of bowel function in 2.8 +/- 0.1 days. The mean postoperative hospital stay was 6.4 +/- 0.4 days. The cardiac and pulmonary status of the patients remained stable during the operation. CONCLUSION: Gasless laparoscopy-assisted colorectal surgery is technically feasible; thus, it provides an alternative means for the performance of minimal-access surgery.

Phase I clinical trial of curcumin, a chemopreventive agent, in patients with high-risk or pre-malignant lesions.

Curcumin (diferuloylmethane), a yellow substance from the root of the plant Curcuma longa Linn., has been demonstrated to inhibit carcinogenesis of murine skin, stomach, intestine and liver. However, the toxicology, pharmacokinetics and biologically effective dose of curcumin in humans have not been reported. This prospective phase-I study evaluated these issues of curcumin in patients with one of the following five high-risk conditions: 1) recently resected urinary bladder cancer; 2) arsenic Bowen's disease of the skin; 3) uterine cervical intraepithelial neoplasm (CIN); 4) oral leucoplakia; and 5) intestinal metaplasia of the stomach. Curcumin was taken orally for 3 months. Biopsy of the lesion sites was done immediately before and 3 months after starting curcumin treament. The starting dose was 500 mg/day. If no toxicity > or = grade II was noted in at least 3 successive patients, the dose was then escalated to another level in the order of 1,000, 2,000, 4,000, 8,000, and 12,000 mg/day. The concentration of curcumin in serum and urine was determined by high pressure liquid chromatography (HPLC). A total of 25 patients were enrolled in this study. There was no treatment-related toxicity up to 8,000 mg/day. Beyond 8,000 mg/day, the bulky volume of the drug was unacceptable to the patients. The serum concentration of curcumin usually peaked at 1 to 2 hours after oral intake of crucumin and gradually declined within 12 hours. The average peak serum concentrations after taking 4,000 mg, 6,000 mg and 8,000 mg of curcumin were 0.51 +/- 0.11 microM, 0.63 +/- 0.06 microM and 1.77 +/- 1.87 microM, respectively. Urinary excretion of curcumin was undetectable. One of 4 patients with CIN and 1 of 7 patients with oral leucoplakia proceeded to develop frank malignancies in spite of curcumin treatment. In contrast, histologic improvement of precancerous lesions was seen in 1 out of 2 patients with recently resected bladder cancer, 2 out of 7 patients of oral leucoplakia, 1 out of 6 patients of intestinal metaplasia of the stomach, I out of 4 patients with CIN and 2 out of 6 patients with Bowen's disease. In conclusion, this study demonstrated that curcumin is not toxic to humans up to 8,000 mg/day when taken by mouth for 3 months. Our results also suggest a biologic effect of curcumin in the chemoprevention of cancer.

Suppression of Fas ligand expression on endothelial cells by arsenite through reactive oxygen species.

Chronic exposure to arsenite is associated with vascular disease, such as arteriosclerosis. However, the cellular mechanisms for vascular disease in response to arsenic are not well known. The present study has demonstrated that arsenite not arsenate decreased the Fas ligand (FasL) expression on ECV304 cells through reactive oxygen species. Incubation of ECV304 cells with arsenite decreased the FasL expression and increased the intracellular peroxide levels. In addition, hydrogen peroxide was found to suppress FasL expression in a dose-dependent manner. The antioxidant, N-acetyl-cysteine, blocked the suppression of FasL expression in response to arsenite. These data suggested that arsenite initiates endothelium dysfunction, at least partly, by suppressing the FasL expression through activating reactive oxygen species sensitive endothelial cell signaling.

Suppression of inducible cyclooxygenase and nitric oxide synthase through activation of peroxisome proliferator-activated receptor-gamma by flavonoids in mouse macrophages.

Peroxisome proliferator-activated receptor (PPAR)gamma transcription factor has been implicated in anti-inflammatory response. Of the compounds tested, apigenin, chrysin, and kaempferol significantly stimulated PPAR gamma transcriptional activity in a transient reporter assay. In addition, these three flavonoids strongly enhanced the inhibition of inducible cyclooxygenase and inducible nitric oxide synthase promoter activities in lipopolysaccharide-activated macrophages which contain the PPAR gamma expression plasmids. However, these three flavonoids exhibited weak PPAR gamma agonist activities in an in vitro competitive binding assay. Limited protease digestion of PPAR gamma suggested these three flavonoids produced a conformational change in PPAR gamma and the conformation differs in the receptor bound to BRL49653 versus these three flavonoids. These results suggested that these three flavonoids might act as allosteric effectors and were able to bind to PPAR gamma and activate it, but its binding site might be different from the natural ligand BRL49653.

Carcinoembryonic antigen in monitoring of response to systemic chemotherapy in patients with metastatic colorectal cancer.

The response to chemotherapy of solid tumors is generally assessed by measuring tumors visualized by imaging. However, the response assessment based on imaging is not always feasible because patients often have disease not measurable by imaging, such as diffuse peritoneal dissemination. We evaluated the correlation between the change on imaging and change in CEA levels for assessing chemotherapeutic response of patients with metastatic colorectal cancer. Between July 1993 and August 1999 we retrospectively examined 136 patients with metastatic colorectal carcinoma, all of whom had measurable lesions. Forty patients received oral tegafur-uracil (300 mg/m2/day) plus folinic acid (60 mg/day) for 4 weeks, repeated every 5 weeks, as the firstline treatment. Another 96 patients received either a weekly intravenous bolus injection of 5-fluorouracil (400 mg/m2) plus folinic acid (20 mg/m2), or an intravenous bolus injection of 5-fluorouracil (425 mg/m2) plus folinic acid (20 mg/m2) for 5 consecutive days every month. Responders, based on CEA assessment, were defined as those with a greater than 50% drop in CEA level for more than 4 weeks. The pretreatment CEA levels were elevated beyond the normal cutoff value in 110 (81%) patients. A response rate of 18.4% (95% CI, 11.9-24.9%), including 8 complete remissions and 17 partial remissions, was achieved according to imaging studies. The response rate assessed by CEA was 25% (34/136). Sixteen responders (47%) based on CEA had no remission on imaging. The sensitivity of change in CEA levels in the prediction of true responders and progressive diseases on imaging were 72% and 81%, respectively. In terms of the positive predictive value, change in CEA levels in the prediction of true responders and progressive disease on imaging were 53% and 85%, respectively. Patients with remarkable falls on CEA levels survived significantly longer than nonresponders (P < 0.001, log-rank test). At follow-up of 48 months the median survival for responders and nonresponders assessed by CEA was 28 months and 13 months, respectively. These data suggest that measurement of CEA levels might be helpful in monitoring chemotherapeutic response when imaging study is unsuitable for assessing the response in clinical practice. Furthermore, measurement of CEA levels may be helpful in determining the prognosis of patients with metastatic colorectal cancer receiving chemotherapy.

Mechanisms of cancer chemoprevention by curcumin.

Curcumin is a major component of the Curcuma species, which is commonly used as a yellow coloring and flavoring agent in foods. Curcumin has shown anti-carcinogenic activity in animals as indicated by its ability to block colon tumor initiation by azoxymethane and skin tumor promotion induced by phorbol ester TPA. Recently, curcumin has been considered by oncologists as a potential third generation cancer chemopreventive agent, and clinical trials using it have been carried out in several laboratories. Curcumin possesses anti-inflammatory activity and is a potent inhibitor of reactive oxygen-generating enzymes, such as lipoxygenase/cyclooxygenase, xanthine dehydrogenase/oxidase and inducible nitric oxide synthase. Curcumin is also a potent inhibitor of protein kinase C, EGF-receptor tyrosine kinase and IkappaB kinase. In addition, curcumin inhibits the activation of NFkappaB and the expression of c-jun, c-fos, c-myc and iNOS. It is proposed that curcumin may suppress tumor promotion by blocking signal transduction pathways in the target cells. Curcumin was first biotransformed to dihydrocurcumin and tetrahydrocurcumin, and these compounds were subsequently convened into monoglucuronide conjugates. The experimental results suggest that curcumin-glucuronide, dihydrocurcumin-glucuronide, tetrahydrocurcumin-glucuronide and tetrahydrocurcumin are major metabolites of curcumin in mice.

Alternative activation of extracellular signal-regulated protein kinases in curcumin and arsenite-induced HSP70 gene expression in human colorectal carcinoma cells.

We have investigated the regulation mechanism of chemical stress-induced HSP70 gene expression in human colorectal carcinoma cells (COLO205 and HT29). Our data show that chemical treatments including sodium arsenite and curcumin, induced significant synthesis of HSP70 and its mRNA. The induced HSP70 gene expression appears to be increased at the transcriptional level. The increase in HSP70 gene expression by both chemicals is associated with an increase in HSF binding to HSE and induction of HSF1 di- or trimerization. Phosphorylation and activation of extracellular signal-regulated proteins (ERK1/2) were detected in sodium arsenite-treated COLO205 and HT29 cells, and the free radical scavenger N-acetyl-L-cysteine (NAC) was able to inhibit this ERK1/2 activation and HSP70 gene expression. MAPK blockade by the specific MEK1 inhibitor (PD98059) decreased the ability of sodium arsenite to increase HSP70 gene expression in a dose-dependent manner along with dephosphorylation of ERK1/2 proteins. In contrast to arsenite treatment, activation of ERK1/2 was not detected in curcumin-treated colorectal carcinoma cells, and NAC and PD98059 did not show any inhibitory effect on HSP70 gene expression induced by curcumin. Overexpression of a dominant negative mutant of mitogen-activated protein kinase kinase kinase 1 (MEKK1-DN) prevents arsenite-induced ERK1/2 phosphorylation and HSP70 protein synthesis. These results indicated that the ERK signaling pathway can participate in HSP70 gene expression induced by the prooxidant sodium arsenite, but not by the antioxidant curcumin.

Induction of apoptosis by garcinol and curcumin through cytochrome c release and activation of caspases in human leukemia HL-60 cells.

Garcinol, a polyisoprenylated benzophenone, was purified from Garcinia indica fruit rind. The effects of garcinol and curcumin on cell viability in human leukemia HL-60 cells were investigated. Garcinol and curcumin displayed strong growth inhibitory effects against human leukemia HL-60 cells, with estimated IC(50) values of 9.42 and 19.5 microM, respectively. Garcinol was able to induce apoptosis in a concentration- and time-dependent manner; however, curcumin was less effective. Treatment with garcinol caused induction of caspase-3/CPP32 activity in a dose- and time-dependent manner, but not caspase-1 activity, and induced the degradation of poly(ADP-ribose) polymerase (PARP). Pretreatment with caspase-3 inhibitor inhibited garcinol-induced DNA fragmentation. Treatment with garcinol (20 microM) caused a rapid loss of mitochondrial transmembrane potential, release of mitochondrial cytochrome c into cytosol, and subsequent induction of procaspase-9 processing. The cleavage of D4-GDI, an abundant hematopoietic cell GDP dissociation inhibitor for the Ras-related Rho family GTPases, occurred simultaneously with the activation of caspase-3 but preceded DNA fragmentation and the morphological changes associated with apoptotic cell death. Of these, Bcl-2, Bad, and Bax were studied. The level of expression of Bcl-2 slightly decreased, while the levels of Bad and Bax were dramatically increased in cells treated with garcinol. These results indicate that garcinol allows caspase-activated deoxyribonuclease to enter the nucleus and degrade chromosomal DNA and induces DFF-45 (DNA fragmentation factor) degradation. It is suggested that garcinol-induced apoptosis is triggered by the release of cytochrome c into the cytosol, procaspase-9 processing, activation of caspase-3 and caspase-2, degradation of PARP, and DNA fragmentation caused by the caspase-activated deoxyribonuclease through the digestion of DFF-45. The induction of apoptosis by garcinol may provide a pivotal mechanism for its cancer chemopreventive action.

Oxidative stress and c-Jun-amino-terminal kinase activation involved in apoptosis of primary astrocytes induced by disulfiram-Cu(2+) complex.

Disulfiram is frequently used in the treatment of alcoholism. In this study, we found that CuCl(2) (1-10 microM), but not other metal ions (Fe(2+), Zn(2+), Pb(2+)), markedly potentiated disulfiram-induced cytotoxicity by 440-fold in primary astrocytes. Thus, the molecular mechanisms of the cytotoxic effects induced by the disulfiram-Cu(2+) complex were explored. The changes in morphology (nuclear condensation and apoptotic body formation) and hypodiploidy of DNA suggested that the disulfiram-Cu(2+) complex induced an apoptotic process. Our studies of the death-signaling pathway reveal that decreased mitochondrial membrane potential, increased free radical production, and depletion of non-protein-thiols (glutathione) were involved. The disulfiram-Cu(2+) complex activated c-Jun-amino-terminal kinase (JNK) and caspase-3 followed by poly (ADP-ribose) polymerase degradation in a time-dependent manner. Moreover, the cellular Cu content was markedly increased and the copper chelator bathocuproine disulfonate abolished all of these cellular events, suggesting that Cu(2+) is essential for death signaling. The antioxidants N-acetylcysteine and vitamin C also inhibited the cytotoxic effect. Thus, we conclude that the disulfiram-Cu(2+) complex induces apoptosis and perhaps necrosis at a late stage mediated by oxidative stress followed by sequential activation of JNK, caspase-3 and poly (ADP-ribose) polymerase degradation. These findings imply that the axonal degeneration and neurotoxicity observed after the chronic administration of disulfiram are perhaps, at least in part, due to the cytotoxic effect of the disulfiram-Cu(2+) complex formed endogenously.