[Tea consumption and cancer: a Mendelian randomization study].

Objective: A Mendelian randomization (MR) analysis was performed to assess the relationship between tea consumption and cancer. Methods: There were 100 639 participants with the information of gene sequencing of whole genome in the China Kadoorie Biobank. After excluding those with cancer at baseline survey, a total of 100 218 participants were included in this study. The baseline information about tea consumption were analyzed, including daily tea consumption or not, cups of daily tea consumption, and grams of daily tea consumption. We used the two-stage least square method to evaluate the associations between three tea consumption variables and incidence of cancer and some subtypes, including stomach cancer, liver and intrahepatic bile ducts cancer, colorectal cancer, tracheobronchial and lung cancer, and female breast cancer. Multivariable MR and analysis only among nondrinkers were used to control the impact of alcohol consumption. Sensitivity analyses were also performed, including inverse variance weighting, weighted median, and MR-Egger. Results: We used 54, 42, and 28 SNPs to construct non-weighted genetic risk scores as instrumental variables for daily tea consumption or not, cups of daily tea consumption, and grams of daily tea consumption, respectively. During an average of (11.4±3.0) years of follow-up, 6 886 cases of cancer were recorded. After adjusting for age, age2, sex, region, array type, and the first 12 genetic principal components, there were no significant associations of three tea consumption variables with the incidence of cancer and cancer subtypes. Compared with non-daily tea drinkers, the HR (95%CI) of daily tea drinkers for cancer and some subtypes, including stomach cancer, liver and intrahepatic bile ducts cancer, colorectal cancer, tracheobronchial and lung cancer, and female breast cancer, are respectively 0.99 (0.78-1.26), 1.17 (0.58-2.36), 0.86 (0.40-1.84), 0.85 (0.42-1.73), 1.39 (0.85-2.26) and 0.63 (0.28-1.38). After controlling the impact of alcohol consumption and performing multiple sensitivity analyses, the results were similar. Conclusion: There is no causal relationship between tea consumption and risk of cancer in population in China.

Antiviral activity of shikonin ester derivative PMM-034 against enterovirus 71 in vitro.

Human enterovirus 71 (EV71) is the major causative agent of hand, foot, and mouth disease (HFMD), particularly in infants and children below 4 years of age. Shikonin is a bioactive compound with anti-inflammatory, antiviral, and antibacterial activities derived from the roots of the Chinese medicinal herb Lithospermum erythrorhizon. This study aimed to examine the antiviral activity of PMM-034, a shikonin ester derivative, against EV71 in rhabdomyosarcoma (RD) cells. Cytotoxicity of PMM-034 on RD cells was determined using WST-1 assay. Dose- and time-dependent effects of PMM-034 on EV71 replication in RD cells were determined using plaque reduction assay. mRNA expression levels of EV71/VP1 and pro-inflammatory cytokines (IL-1ß, IL-6, IL-8, and TNF-α) were determined by real-time RT-PCR, and EV71/VP1 and phospho-p65 protein expressions were determined by western blot analysis. PMM-034 exhibited only weak cytotoxicity against RD cells. However, PMM-034 exhibited significant antiviral activity against EV71 in RD cells with 50% inhibitory concentration of 2.31 µg/mL. The VP1 mRNA and protein levels were significantly reduced in cells treated with PMM-034. Furthermore, relative mRNA expression levels of IL-1ß, IL-6, IL-8, and TNF-α significantly decreased in the cells treated with PMM-034, while the phospho-p65 protein expression was also significantly lower in the treated cells. These results indicated that PMM-034 suppressed the expressions of pro-inflammatory cytokines in RD cells, exhibiting antiviral activity against EV71, as evidenced by the reduced VP1 mRNA and protein levels in PMM-034-treated cells. Thus, PMM-034 is a promising candidate for further development as an EV71 inhibitor.

[An analysis of spectrum of allergen sensitization among patients with tree pollen allergy in Beijing].

Objective:To explore the feature of allergic sensitization of tree pollen and the relationship between tree pollen and airborne pollen in Beijing. Method:Two thousand five hundred and twenty-one patients with allergic diseases were enrolled between January to July in 2017 in an allergy department. All patients received SPT of 8 kinds of tree pollen. Result:①The overall tree pollen positive rate of SPT was 49.3% (1 244/2 521). Male had a higher positive rate than female (56.4% vs 45.0%, P<0.01) while child group had a higher positive rate than adult group (55.9% vs 48.6%, P<0.05). The poly sensitization was observed with a rate of 26.3%, take the percentage of 53.3% of the overall positive subjects. ②The highest sensitization rate was found in March (62.2%), followed by May (52.1%), the lowest was January (27.9%). There was a positive correlation between tree pollen sensitization rate and pollen count of each month (r=0.929, P<0.01). ③Cypress had the highest positive rate (34.6%), followed by poplar (21.3%), the lowest was sophora (13.3%). The sensitization rate was positively correlated with pollen count among different tree pollen (r=0.714, P<0.05). Conclusion:The main allergic pollen in Beijing was cypress. The tree pollen sensitization showed a seasonality with the peak in March. The sensitization status was tightly associated with the amount of airborne pollen.

Antiviral activity of shikonin ester derivative PMM-034 against enterovirus 71 in vitro

Human enterovirus 71 (EV71) is the major causative agent of hand, foot, and mouth disease (HFMD), particularly in infants and children below 4 years of age. Shikonin is a bioactive compound with anti-inflammatory, antiviral, and antibacterial activities derived from the roots of the Chinese medicinal herb Lithospermum erythrorhizon. This study aimed to examine the antiviral activity of PMM-034, a shikonin ester derivative, against EV71 in rhabdomyosarcoma (RD) cells. Cytotoxicity of PMM-034 on RD cells was determined using WST-1 assay. Dose- and time-dependent effects of PMM-034 on EV71 replication in RD cells were determined using plaque reduction assay. mRNA expression levels of EV71/VP1 and pro-inflammatory cytokines (IL-1β, IL-6, IL-8, and TNF-α) were determined by real-time RT-PCR, and EV71/VP1 and phospho-p65 protein expressions were determined by western blot analysis. PMM-034 exhibited only weak cytotoxicity against RD cells. However, PMM-034 exhibited significant antiviral activity against EV71 in RD cells with 50% inhibitory concentration of 2.31 μg/mL. The VP1 mRNA and protein levels were significantly reduced in cells treated with PMM-034. Furthermore, relative mRNA expression levels of IL-1β, IL-6, IL-8, and TNF-α significantly decreased in the cells treated with PMM-034, while the phospho-p65 protein expression was also significantly lower in the treated cells. These results indicated that PMM-034 suppressed the expressions of pro-inflammatory cytokines in RD cells, exhibiting antiviral activity against EV71, as evidenced by the reduced VP1 mRNA and protein levels in PMM-034-treated cells. Thus, PMM-034 is a promising candidate for further development as an EV71 inhibitor.

[Aluminuminduced impairment in primary cultured rat choroid plexus epithelial cells].

OBJECTIVE: To investigate the impairment in primary cultured rat choroid plexus epithelial cells (CPECs)induced by aluminum. METHODS: The choroid plexus isolated from Sprague-Dawley rats 14 days old was cut into pieces and digested by trypsin in the sterile area. The obtained single cells were cultured in DMEM with 1% epidermal growth factor and 20% fetal calf serum. Five days later, immunohistochemistry with anti-transthyretin antibody was used to identify the purity of cultured cells. The well-grown cells were treated with aluminum lactate at different concentrations (0, 100, 400, and 1 600 µmol/L for control, lowdose, mediumdose, and highdose groups). Fortyeight hours later, the cell viability, apoptotic rate, level of reactive oxygen species (ROS), and activity of superoxide dismutase (SOD)were measured in each group to evaluate the impairment in primary cultured rat CPECs by aluminum. RESULTS: More than 95% of the cultured cells were identified as CPECs. The medium-and high-dose groups had significantly lower cell viability than the control group(86.74%±4.03% vs 100%, P<0.01; 81.90%±9.17% vs 100%, P<0.01). The high-dose group had significantly lower cell viability than the lowdose group (81.90%±9.17% vs 92.92%±8.81%, P<0.01). The medium-and high-dose groups had significantly higher apoptotic rates than the control group (7.26%±0.99% vs 1.29%±0.03%, P<0.01; 22.25%±1.55% vs 1.29%±0.03%, P<0.01)and the low-dose group (7.26%±0.99% vs 1.68%±0.27%, P<0.01; 22.25%±1.55% vs 1.68%±0.27%, P<0.01). The high-dose group had a significantly higher apoptotic rate than the medium-dose group (22.25%±1.55% vs 7.26%±0.99%, P<0.01). The mediumand high-dose groups had significantly higher fluorescence intensity of ROS than the control group (22.23%±0.41% vs 17.24%±0.09%, P<0.05; 25.10%±1.13% vs 17.24%±0.09%, P<0.05)and the lowdose group (22.23%±0.41% vs 18.31%±0.21%, P<0.05; 25.10%±1.13% vs 18.31%±0.21%, P<0.05). The highdose group had significantly higher fluorescence intensity of ROS than the mediumdose group (25.10%±1.13% vs 22.23%±0.41%, P< 0.05). The low-, medium-and high-dose groups had significantly lower SOD activity than the control group[(28.65±0.74)U/g Hb vs (37.35±1.05)U/g Hb, P<0.05; (22.75±1.94)U/g Hb vs (37.35±1.05)U/g Hb, P<0.05; (13.29±0.64)U/g Hb vs(37.35±1.05)U/g Hb, P<0.05]. The medium-and high-dose groups had significantly lower SOD activity than the low-dose group[(22.75±1.94)U/g Hb vs(28.65±0.74)U/g Hb, P<0.05; (13.29±0.64)U/g Hb vs (28.65±0.74)U/g Hb, P<0.05], while the high-dose group had had significantly lower SOD activity than the medium-dose group[(13.29±0.64)U/g Hb vs (22.75±1.94)U/g Hb, P<0.05]. There were no significant differences in cell viability, apoptotic rate, level of ROS, or activity of SOD between any other two groups (P>0.05). CONCLUSION: Aluminum lactate may induce impairment in primary cultured rat CPECs. It reduces the cell viability, elevates the apoptotic rate, and causes oxidative stress.

Cloning and sequence analysis of the Blumea balsamifera DC farnesyl diphosphate synthase gene.

Blumea balsamifera DC is a member of the Compositae family and is frequently used as traditional Chinese medicine. Blumea balsamifera is rich in monoterpenes, which possess a variety of pharmacological activities, such as antioxidant, anti-bacteria, and anti-viral activities. Farnesyl diphosphate synthase (FPS) is a key enzyme in the biosynthetic pathway of terpenes, playing an important regulatory role in plant growth, such as resistance and secondary metabolism. Based on the conserved oligo amino acid residues of published FPS genes from other higher plant species, a cDNA sequence, designated BbFPS, was isolated from B. balsamifera DC using polymerase chain reaction. The clones were an average of 1.6 kb and contained an open reading frame that predicted a polypeptide of 342 amino acids with 89.07% identity to FPS from other plants. The deduced amino acid sequence was dominated by hydrophobic regions and contained 2 highly conserved DDxxD motifs that are essential for proper functioning of FPS. Phylogenetic analysis indicated that FPS grouped with other composite families. Prediction of secondary structure and subcellular localization suggested that alpha helices made up 70% of the amino acids of the sequence.

Linezolid in the treatment of extensively drug-resistant tuberculosis.

PURPOSE: To investigate the linezolid (LZD) treatment outcome and correlation between in vitro susceptibility to LZD and clinical outcome. METHODS: We retrospectively reviewed records of tuberculosis (TB) patients who received treatment with linezolid between March 2012 and February 2013. RESULTS: A total of 43 extensively drug-resistant (XDR) TB patients identified by drug susceptibility testing were enrolled in this study, including 15 (34.9 %) received LZD as part of individualized treatment regimens. Among the 43 XDR TB patients, 15 patients (34.9 %) obtained favorable clinical outcome, including 9 (60.0 %) from LZD group and 6 (21.4 %) from control group without LZD. Statistical analysis revealed that the percentage of favorable outcomes of LZD group was significantly higher than that of control group (P = 0.011). Furthermore, we analyzed the LZD minimum inhibitory concentrations of Mycobacterium tuberculosis (MTB) isolates from patients in LZD group and identified 4 (26.7 %) resistant to LZD. All of the patients with LZD resistance harbored adverse clinical outcome, while most of the patients infected with LZD sensitive MTB harbored favorable clinical outcome (81.8 %, 9/11). Statistical analysis revealed that the percentage of favorable outcome among the patients with LZD resistance was statistically lower than that among the LZD susceptible group (P = 0.011). CONCLUSION: This study demonstrates that linezolid has efficacy against XDR pulmonary TB patients, even in shorter duration of administration. The XDR TB patients infected with LZD-resistant isolates were more likely to obtain the adverse clinical outcome under the treatment of regimen containing LZD.

Genetic diversity of the Chinese traditional herb Blumea balsamifera (Asteraceae) based on AFLP markers.

Blumea balsamifera is a commercially important medicinal herb in China and other parts of Asia. It is used to produce borneol. This plant grows in the wild, but resources have diminished greatly in recent years. We examined the genetic diversity of this species to help develop conservation strategies; 35 plants from five provinces were analyzed using AFLPs. Eight AFLP primer combinations generated 1367 fragments, giving a mean of 172 fragments per primer combination. Polymorphism in the germplasm analysis was found for 1360 (99.48%) of the fragments, of which 264 (19.27%) fragments were unique (accession specific) and 423 (25.33%) of the fragments were rare (present in less than 10% of the accessions). The polymorphic fragments were used to group the accessions in a UPGMA phenogram. Most grouping was geographical. In general, accessions coming from Guizhou and Guangxi showed higher diversities as these accessions were scattered in different groups. The genetic distance estimated by Jaccard similarity coefficient index showed low variability among genotypes (coefficient value ranged from 0.60 to 0.95). More attention should be given to the study and conservation of the biodiversity of this economically important genus.

Diagnostic dilemma: treatment outcomes of tuberculosis patients with inconsistent rifampicin susceptibility.

OBJECTIVE: A retrospective clinical trial to evaluate treatment outcomes in adults with smear-positive tuberculosis (TB) and discordant rifampicin (RMP) resistance results. DESIGN: A total of 2156 smear-positive TB patients underwent both conventional and Genechip drug susceptibility testing (DST) for RMP resistance. All 49 patients with discordant results treated with either a first-line or second-line regimen were analysed. RESULTS: Of 30 Type I cases (Genechip-resistant, conventional DST-susceptible) receiving the first-line regimen, 4 had a favourable outcome and 5 failed treatment. The 21 remaining Type I cases were treated with the second-line regimen, of whom 18 had a favourable outcome. Second-line regimen thus resulted in significantly more favourable outcomes than first-line treatment (P = 0.032). Among Type II cases (Genechip-susceptible, conventional DST-resistant), 13/19 received the first-line regimen, and 7 had a favourable outcome. The six Type II cases treated with the second-line regimen all had favourable outcomes. CONCLUSION: Patients with discordant RMP DST results who receive second-line regimens may have a better clinical response than those treated with the first-line regimen. Patients infected with fluoroquinolone-resistant Mycobacterium tuberculosis strains were observed to have a significantly higher treatment failure rate.

Pharmacokinetics of 23-epi-26-deoxyactein in women after oral administration of a standardized extract of black cohosh.

Dietary supplements containing black cohosh are alternatives to conventional hormone replacement therapy in menopause. This study investigates the maximum tolerated dose of a 75% ethanol extract of black cohosh and determines the pharmacokinetics of one of its most abundant triterpene glycosides, 23-epi-26-deoxyactein. Single doses of black cohosh extract containing 1.4, 2.8, or 5.6 mg of 23-epi-26-deoxyactein were administered to 15 healthy, menopausal women. Serial blood samples and 24-h urine samples were obtained; blood chemistry, hormonal levels, and 23-epi-26-deoxyactein levels were determined. No acute toxicity or estrogenic hormone effects were observed. Pharmacokinetic analyses of 23-epi-26-deoxyactein in sera indicated that the maximum concentration and area under the curve increased proportionately with dosage, and that the half-life was ~2 h for all dosages. Less than 0.01% of the 23-epi-26-deoxyactein was recovered in urine 24 h after administration. No phase I or phase II metabolites were observed either in clinical specimens or in vitro.

The influence of copper concentration and source on ileal microbiota.

Copper is normally supplemented in poultry diets as a growth promotant and antimicrobial. However, there are conflicting reports about the growth benefits and little information about how Cu affects the microbiota in the intestinal tract of poultry. Therefore, in vitro and in vivo experiments were conducted with broilers to determine the effects of Cu source and supplementation on ileal microbiota. The influence of Cu on growth of lactobacilli and Escherichia coli in media inoculated with ileal contents was determined in the first study. When Cu sulfate pentahydrate was supplemented to the cultures, quadratic increases in lactobacilli to graded concentrations of Cu up to 125 mg/kg and quadratic decreases in E. coli up to 250 mg/kg of Cu were observed after 24 h of incubation at 37 degrees C. However, when tribasic Cu chloride (TBCC) was supplemented, neither linear nor quadratic responses to graded concentrations of dietary Cu were observed on number of lactobacilli or number of E. coli. The effects of Cu and Cu source on ileal microbiota and growth performance in broiler chickens were determined in the second study. Bird performance was not affected by Cu source or concentration. The bacterial culture enumeration results revealed that supplementation with 187.5 mg/kg of Cu from Cu sulfate pentahydrate and TBCC had no effect on number of ileal lactobacilli of birds. The denaturing gradient gel electrophoresis analyses of ileal microbial communities revealed that neither Cu supplementation nor source had effects on the number of bacterial species predominant in the ileal digesta or associated with the ileal mucosa. Supplementation with TBCC supplementation significantly increased the similarity coefficients of microbiota in the ileal mucosa compared with cross-products of all individuals. This suggests that TBCC may alter the intestinal microbiota, yet this shift had no effect on bird performance.

East meets West in the search for Alzheimer's therapeutics - novel dimeric inhibitors from tacrine and huperzine A.

Alzheimer's disease (AD) is linked to cholinergic deficiency and the overactivation of glutamate receptors. The acetylcholinesterase (AChE) inhibition treatment approach has produced the most encouraging results in clinical practice, and memantine, a moderate antagonist of N-methyl-D-aspartate (NMDA) receptors, has been approved for treating AD. However, AChE inhibitors have limited success as they only improve memory in mild dementia but cannot stop the process of neurodegeneration; while memantine possesses neuroprotective effects only with a little ability in memory enhancement. There has been a major rush among neuroscience research institutions and pharmaceutical firms worldwide to search for safer and more effective therapeutic agents for AD. The novel dimers, derived from tacrine and the fragment of huperzine A (HA'), have been demonstrated to be potent and selective reversible inhibitors of AChE. Bis(7)-tacrine, bis(12)-hupyridone (E12E) and HA'(10)-tacrine, are representatives of three series of novel dimers. According to the preclinical studies, these compounds have been shown to have low toxicity and high efficacy for improving cognitive deficits in several animal models. More interestingly, bis(7)-tacrine, similar to memantine, prevents glutamate-induced neurotoxicity by moderately blocking glutamate receptor NMDA subtype. Furthermore, bis(7)-tacrine, as well as E12E, possesses multiple neuroprotective effects in vitro and in vivo. Taking together, these dimeric AChE inhibitors, especially bis(7)-tacrine, E12E and HA'(10)-tacrine, may provide beneficial effects in AD and other neurodegenerative diseases.

Effects of dietary copper supplementation and copper source on digesta pH, calcium, zinc, and copper complex size in the gastrointestinal tract of the broiler chicken.

An experiment was conducted to study the effects of high dietary Cu and Cu source on the pH of digesta from the gizzard, duodenum + jejunum, ileum, and complex size of Ca, Zn, and Cu in the duodenum + jejunum digesta of broiler chickens. Ross 308 male broiler chicks were randomly assigned to 32 cages and fed 1 of 4 treatments: control, 250 ppm Cu from sulfate, 250 ppm Cu from lysinate, and 250 ppm tribasic Cu from chloride from 15 to 21 d of age. Copper supplementation and Cu source had no effects on pH of gizzard or duodenum + jejunum contents. Copper supplementation, however, increased the pH of the ileal contents (P < 0.05) but was not affected by Cu source. Neither Cu supplementation nor Cu source had significant effects on the solubility of Ca in the duodenum + jejunum contents, and the portions of Ca existing in different soluble complex sizes: >100,000, 100,000 to 30,000, 30,000 to 5,000, and <5,000 molecular weight (MW) in the duodenum + jejunum digesta. About 80% of soluble Ca, Cu, and Zn was associated with either large complexes (>100,000 MW) or small complexes (<5,000 MW). The solubility of supplemental Cu in digesta was from 59 to 61% (P < 0.05), but solubility was not affected by Cu source. No effects on portions of Cu existing in different sizes of complexes in the supernatant were noted. Copper lysinate decreased the Zn solubility in the digesta (P < 0.05), but Cu sulfate and tribasic Cu chloride supplementation did not. Copper supplementation increased (P < 0.05) the percentage of Zn associated with large complexes (>100,000 MW) and decreased (P < 0.05) the percentage of Zn associated with small complexes (<5,000 MW; P < 0.05), thereby suggesting an antagonism between Cu and Zn.

Schisandrin B protects against tacrine- and bis(7)-tacrine-induced hepatotoxicity and enhances cognitive function in mice.

Intragastric administration (100-200 micromol/kg) of tacrine (THA) or bis(7)-THA could cause an acute and dose-dependent increase in plasma alanine aminotransferases activity in mice at 6 h after the drug administration. The increase in plasma enzyme activity was associated with an increase in hepatic malondialdehyde level, an indirect index of oxidative tissue damage. Pretreating mice with schisandrin B (Sch B), an active dibenzocyclooctadiene derivative isolated from the fruit of Schisandra chinensis, at a daily dose of 0.125-0.5 mmol/kg for 3 days protected against the THA/bis(7)-THA induced hepatic oxidative damage in a dose-dependent manner. Sch B treatment (0.025-0.5 mmol/kg/day x 5) also enhanced the passive avoidance-response in mice as assessed by the step-through task experiment. The ensemble of results suggests that Sch B may be useful for reducing the potential hepatotoxicity of THA/bis(7)-THA in anti-Alzheimer's therapy.

Discovery of a new inhibitor lead of adenovirus proteinase: steps toward selective, irreversible inhibitors of cysteine proteinases.

Using the computer docking program EUDOC, in silico screening of a chemical database for inhibitors of human adenovirus cysteine proteinase (hAVCP) identified 2,4,5,7-tetranitro-9-fluorenone that selectively and irreversibly inhibits hAVCP in a two-step reaction: reversible binding (Ki = 3.09 microM) followed by irreversible inhibition (ki = 0.006 s(-1)). The reversible binding is due to molecular complementarity between the inhibitor and the active site of hAVCP, which confers the selectivity of the inhibitor. The irreversible inhibition is due to substitution of a nitro group of the inhibitor by the nearby Cys122 in the active site of hAVCP. These findings suggest a new approach to selective, irreversible inhibitors of cysteine proteinases involved in normal and abnormal physiological processes ranging from embryogenesis to apoptosis and pathogen invasions.

Synthesis and evaluation of novel aldose reductase inhibitors: Effects on lens protein kinase Cgamma.

PURPOSE: To synthesize novel aldose reductase inhibitors (ARI) that will normalize losses in protein kinase Cgamma (PKCgamma) observed during diabetes and galactosemia. METHODS: ARI were synthesized as tricyclic pyrones 1-6 (HAR-1 through HAR-6) from 3-methyl-1H,7H-5a,6,8,9-tetrahydro-1-oxopyrano[4,3-b][1]benzopyran and (5aS,7S)-7-isopropenyl-3-methyl-1H,7H-5a,6,8,9-tetrahydro-1-oxopyrano[4,3-b][1]benzopyran and were tested by inhibition of aldose reductase enzyme activity in vitro and by inhibition of polyol formation in lens epithelial cells in culture. Identified compounds were further tested in galactosemic rat lens in vivo for (a) normalized PKCgamma levels by Western blot, (b) reduction of phosphorylation of the gap junction protein Cx46 by analyses of co-immunoprecipitated proteins, and (c) by normalization of gap junction activity as measured by dye transfer. RESULTS: HAR-1 (1H,7H-5a,6,8,9-tetrahydro-1-oxopyrano[4,3-b][1]benzopyran-3-acetic acid) was identified as an ARI with IC50 for aldose reductase inhibition at 2 nM. Polyol accumulation in lens epithelial cells was reduced by 80% at 10 microM. Rats fed 40% galactose for 9 days had an 80% reduction in PKCgamma levels which were normalized by HAR-1 at 100 mg/kg/day, fed orally. Phosphorylation of Cx46 was increased by 50% and this was normalized in HAR-1 treated rats (6 day treatment). Gap junction activity of galactosemic rats was reduced by 55% and this was normalized by HAR-1 in six day-treated rats. CONCLUSIONS: HAR-1 is a novel ARI which normalized losses of PKCgamma, changes in Cx46 phosphorylation, and gap junction activity.

Insulin-like growth factor I receptor antagonism augments response to chemoradiation therapy in colon cancer cells.

BACKGROUND: Colorectal cancer remains one of the most prevalent malignancies in the United States. Improvement in local disease control is seen when 5-fluorouracil (5-FU) is used in combination with pelvic irradiation for rectal adenocarcinoma. The frequent overexpression of insulin-like growth factor I receptor (IGF-I-R) in rectal adenocarcinoma suggests that inhibition of the signal transduction pathway may be a novel approach to enhance tumor response. This investigation seeks to define the role of IGF-I-R antagonism, using monoclonal antibody alpha-IR3, in augmenting cytotoxicity to adjuvant chemoradiation therapy for adenocarcinoma of the rectum. MATERIALS AND METHODS: SW 480 colon cancer cells were cultured to semiconfluent conditions with dose titrations performed for 5-FU to determine that the IC(50) (inhibitory concentration of 50% of the cells) was 0.5 microg/ml. The IC(50) for 5-FU was reassessed in the presence of IGF-I. Experimental groups included colon cancer cells combined with 5-FU; 6-MeV external beam radiation (100-500 cGy); and alpha-IR-3. RESULTS: The addition of 100 ng/ml IGF-I 1 h prior to 5-FU or radiation significantly blunted the expected cytotoxicity, resulting in a 10-fold increase in the IC(50) (from 0.5 to 5 microg/ml). Receptor antagonism using the monoclonal antibody alpha-IR-3 (100-400 ng/ml) produced a dose-dependent increase in cytotoxicity compared with 5-FU alone. The addition of radiation produced synergistic amplification of this response. CONCLUSIONS: IGF-I-R activation blocks the expected cytotoxic effects of 5-FU and external beam radiation. Receptor antagonism increased the cytotoxic response of chemoradiation therapy. These data suggest the utility of inhibiting IGF-I-R signal transduction in the treatment of rectal adenocarcinoma.

Atomic levers control pyranose ring conformations.

Atomic force microscope manipulations of single polysaccharide molecules have recently expanded conformational chemistry to include force-driven transitions between the chair and boat conformers of the pyranose ring structure. We now expand these observations to include chair inversion, a common phenomenon in the conformational chemistry of six-membered ring molecules. We demonstrate that by stretching single pectin molecules (1 --> 4-linked alpha-D-galactouronic acid polymer), we could change the pyranose ring conformation from a chair to a boat and then to an inverted chair in a clearly resolved two-step conversion: 4C1 right arrow over left arrow boat right arrow over left arrow 1C4. The two-step extension of the distance between the glycosidic oxygen atoms O1 and O4 determined by atomic force microscope manipulations is corroborated by ab initio calculations of the increase in length of the residue vector O1O4 on chair inversion. We postulate that this conformational change results from the torque generated by the glycosidic bonds when a force is applied to the pectin molecule. Hence, the glycosidic bonds act as mechanical levers, driving the conformational transitions of the pyranose ring. When the glycosidic bonds are equatorial (e), the torque is zero, causing no conformational change. However, when the glycosidic bond is axial (a), torque is generated, causing a rotation around C---C bonds and a conformational change. This hypothesis readily predicts the number of transitions observed in pyranose monomers with 1a-4a linkages (two), 1a-4e (one), and 1e-4e (none). Our results demonstrate single-molecule mechanochemistry with the capability of resolving complex conformational transitions.

Bis(7)-tacrine, a novel dimeric AChE inhibitor, is a potent GABA(A) receptor antagonist.

Heptylene-linked bis-(9-amino-1,2,3,4-tetrahydroacridine) (bis(7)-tacrine) is a potential palliative therapeutic agent for Alzheimer's disease (AD), on the basis of its superior acetylcholinesterase (AChE) inhibition and memory-enhancing potency relative to tacrine. In this study we report that bis(7)-tacrine exhibits a potentially complementary central nervous system action, antagonism of GABA(A) receptor function. Bis(7)-tacrine displaced [3H]muscimol from rat brain membranes with an apparent Ki of 6.0 microM; tacrine and physostigmine were shown to be 18 and 170 times less potent, respectively. In whole-cell patch-clamp recordings, bis(7)-tacrine inhibited GABA-induced inward current with an IC50 of 5.6 microM, and shifted the GABA concentration-response curve to the right in a parallel manner. These results suggest that bis(7)-tacrine is a competitive antagonist of the GABA(A) receptor.

Dual-site binding of bivalent 4-aminopyridine- and 4-aminoquinoline-based AChE inhibitors: contribution of the hydrophobic alkylene tether to monomer and dimer affinities.

Three series of 4-aminopyridine-and 4-aminoquinoline based symmetrical bivalent acetylcholinesterase (AChE) inhibitors were prepared and compared to previously synthesized dimers of 9-amino-1,2,3,4-tetrahydroacridine (tacrine). In each case significant, tether length-dependent increases in AChE inhibition potency and selectivity (up to 3000-fold) were observed relative to the corresponding monomer, indicating dual-site binding of these inhibitors to AChE. Assay of the corresponding alkylated monomers revealed that the alkylene tether played at least two complementary roles in the dimer series. In addition to reducing the entropy loss that occurs on binding both monomeric units of the dimer, the alkylene tether can also significantly improve potency through hydrophobic effects.