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Pancreatic tumors are hypovascular, which leads to a poor nutrient supply to support the aggressively proliferating tumor cells. However, human pancreatic cancer cells have extreme resistance to nutrition starvation, which enables them to survive under severe metabolic stress conditions within the tumor microenvironment, a phenomenon known as "austerity" in cancer biology. Discovering agents which can preferentially inhibit the cancer cells' ability to tolerate starvation conditions represents a new generation of anticancer agents. In this study, geranyl 2,4-dihydroxy-6-phenethylbenzoate (GDP), isolated from Boesenbergia pandurata rhizomes, exhibited potent preferential cytotoxicity against PANC-1 human pancreatic cancer cells under nutrition starvation conditions. GDP also possessed PANC-1 cell migration and colony formation inhibitory activities under normal nutrient-rich conditions. Mechanistically, GDP inhibited PI3K/Akt/mTOR/autophagy survival signaling pathway, leading to selective PANC-1 cancer cell death under the nutrition starvation condition. Therefore, GDP is a promising anti-austerity agent for drug development against pancreatic cancer.

It has been reported that vitamin D regulates the immune system. However, whether vitamin D repletion modulates inflammatory responses in lymphocytes from dialysis patients is unclear. In the clinical trial, thirty-two (32) dialysis patients with 25 vitamin D ≤ 20ng/mL were randomized to receive either supplementation of cholecalciferol 100,000 UI/week/3 months (16 patients) or placebo (16 patients). In the in vitro study, B and T lymphocytes from 12 healthy volunteers (HV) were incubated with or without uremic serum in the presence or absence of 25 or 1,25 vitamin D. We evaluated the intracellular expression of IL-6, IFN-γ TLR7, TLR9, VDR, CYP27b1 and CYP24a1 by flow cytometry. We observed a reduction in the expression of TLR7, TLR9, INF-γ and CYP24a1 and an increase in VDR and CYP27b1 expression in patients which were supplemented with cholecalciferol, whereas no differences were found in the placebo group. Uremic serum increased the intracellular expression of IL-6, IFN-γ, TLR7, TLR9, VDR, CYP27b1 and CYP24a1. Treatment with 25 or 1,25 vitamin D decreased IL-6 and TLR9. CYP24a1 silencing plus treatment with 25 and/or 1,25 vitamin D had an additional reduction effect on IL-6, IFN-γ, TLR7 and TLR9 expression. This is the first study showing that cholecalciferol repletion has an anti-inflammatory effect and improves vitamin D intracellular regulatory enzymes on lymphocytes from dialysis patients.

We compared recombinant human chymase expressed in Escherichia coli with human chymase purified from vascular tissues. The recombinant chymase, the structure of which was NH2-enterokinase cleavage site-chymase-COOH, was expressed in Escherichia coli and then was solubilized and renatured. The protein did not have a chymase activity, but gained this activity after the cleavage of the N-terminal site by enterokinase. The enzyme was purified by heparin affinity and gel filtration columns. The N-terminal sequence of the protein was identical to the sequence for human chymase. The molecular weights of the recombinant chymase and chymase purified from human vascular tissues were 26 and 30 kDa, respectively, and the 4 kDa difference was thought to be due to the presence or absence of glycan. The optimum pH of the recombinant enzyme activity was between 7.5 and 9.0. The activity of the recombinant enzyme was inhibited by chymostatin, soybean trypsin inhibitor and phenylmethylsulfonyl fluoride, but not by ethylenediaminetetraacetic acid and aprotinin. This enzyme cleaved specifically the Phe8-His9 bond of angiotensin (Ang) I to form Ang II and that of big endothelin (ET)-l to form ET-1-(1-31). These findings demonstrated that the enzymatic characteristics of the recombinant enzyme were identical to that of native human chymase.

Helicobacter pylori (Hp) has strong urease activity and produces a large amount of ammonia in the stomach. In animal studies, ammonia was shown to accelerate cell kinetics of gastric mucosa, and long-term exposure of the stomach to ammonia leads to mucosal atrophy. To understand this process, we examined the effects of ammonia on the growth and cell cycle progression of human gastric cancer cell lines (HGC-27, MKN1, MKN45) using flow-cytometric analysis. In each cell line, ammonia inhibited the cell growth in a dose-dependent manner and caused significant accumulation of S-phase cells at a cytostatic dose. DNA synthesis of HGC-27 cells treated with ammonia was also suppressed to about 50% of that of the untreated cells. Similar effects were observed on addition of ammonium chloride at the same concentration, while adjusting the pH of the media with NaOH alone to that with the cytostatic dose of ammonia did not affect the cell cycle progression. These observations indicate that ammonia induces S-phase arrest in gastric cells independently of pH.

The clinical benefits of high-performance membrane (HPM) dialyzers have often been reported since the advent of the synthetic polyacrylonitrile dialysis membrane. HPMs, which have high water permeability, eliminate a wide spectrum of uremic toxins, and offer excellent biocompatibility, are now essential for hemodialysis, hemofiltration, and hemodiafiltration. For HPMs whose mean pore size is enlarged to secure better dialysis membrane performance, however, the dialyzing fluid must be highly purified to prevent contamination. Blood purification therapies using highly purified dialyzing fluid and HPM are expected to improve the prognosis of future patients undergoing dialysis.

Quercetin, a flavonoid, is found in many plants, including edible fruits and vegetables. We examined the effects on cell growth of human malignant cells derived from the gastrointestinal tract and on cell cycle progression. Quercetin markedly inhibited the growth of human gastric cancer cells and the IC50 value was 32-55 microM. DNA synthesis was suppressed to 14% of the control level by the treatment with 70 microM quercetin for 2 days. Furthermore, quercetin blocked cell progression from the G1 to the S phase.

Using a human neuroblastoma cell line GOTO, the effects of delta 12-prostaglandin (PG) J2 on the modulation of cell cycle progression and protein synthesis were examined in comparison with those caused by heat shock (HS). delta 12-PGJ2 induced G1 arrest, the peak of which was obtained at 24 h and continued for 72 h. HS was found to induce G1 arrest earlier than delta 12-PGJ2. Furthermore, sequential HS could maintain G1 arrest. delta 12-PGJ2 induced the synthesis of several heat shock proteins (HSPs) in a manner similar to HS. Using immunoblot analysis, HSP72 was detected prior to inducing G1 arrest and accumulated during the subsequent 72h. The content of HSP72 induced by HS also correlated well with the induction, release, and maintenance of G1 arrest. In addition, both delta 12-PGJ2 and HS induced HSP72 mRNA and simultaneously suppressed N-myc mRNA expression. These results suggest that delta 12-PGJ2 and HS regulate cell cycle progression of GOTO cells via similar mechanisms.

Cells exposed to several forms of stress, such as heat shock, transiently synthesize a group of proteins called heat shock proteins (hsps). Although many stressors other than heat shock are known to induce hsps, inhibitors of hsp expression have never been reported. Here we show that quercetiri and several other flavonoids inhibit the synthesis of hsps induced by heat shock in two human cell lines, Hela cells and COLO320 DM cells. Quercetin inhibited the induction of hsp70 at the level of mRNA accumulation. This is the first report to describe the inhibition of hsp expression by reagents.

Quercetin inhibits growth of COLO320 DM cells, derived from a human colon cancer. The inhibitory effect is partially reversible when quercetin is removed from the culture medium. Flow cytometric analysis has revealed that quercetin causes perturbation of the cell cycle, inducing a frozen cell-cycle pattern and a block at the G1/S boundary. The synthesis of a 17-kDa protein was specifically inhibited by the addition of quercetin, and recovered when the cells at the G1/S boundary progressed into S-phase after the removal of quercetin from the culture medium. Furthermore, using synchronized cells obtained by centrifugal elutriation, we have shown that the rate of synthesis of a 17-kDa protein was low in G1, and high in S-phase of the cell cycle. Thus, this protein appears to be cell-cycle-related.

Detailed enzymatic properties of the ureido ring synthetase purified from Pseudomonas graveolens were investigated. Nucleotide specificity studies indicated that CTP, UTP, GTP, and ITP were each tenth to one-fifth as active as ATP. The effect of substrate concentration was examined. The Km values for 7, 8-diaminopelargonic acid, biotin diaminocarboxylic acid, NaHCO3, ATP, and MgCl2 were 1×10-4M, 4×10-5M, 1×10-2M, 5×10-5M, and 3×10-3M, respectively. It was elucidated that only ADP was produced from ATP in both the reaction of desthiobiotin synthesis from 7, 8-diaminopelargonic acid and biotin synthesis from biotin diaminocarboxylic acid. The reaction was remarkably inhibited by Ni2+, Cd2+, Cu2+, Ag+, and As3+, while Mn2+ remarkably enhanced the enzyme reaction. The reaction was remarkably inhibited by metal-chelating reagents. It was elucidated that ADP had a competitively inhibiting effect on this enzyme reaction. 7, 8-Diaminopelargonic acid, which is the substrate for the desthiobiotin synthesis, competitively inhibited the biotin synthesis from biotin diaminocarboxylic acid. The stoichiometry of the desthiobiotin synthesis indicated that the formation ratio of desthiobiotin to ADP was 1 to 1.

Effects of cyclopentenone prostaglandins, delta 12-prostaglandin (PG) J2 and PGA2 on the expression of N-myc in relation to the effects on cell cycle progression were investigated using human neuroblastoma cell line GOTO. Both PGs suppressed N-myc expression within several hours prior to inducing G1 arrest. The N-myc suppression with delta 12-PGJ2 was continued but with PGA2 it was gradually released, followed by the release of G1 arrest. These results suggest that delta 12-PGJ2 and PGA2 inhibit cell cycle progression in strong association with N-myc suppression and delta 12-PGJ2 is more potent and has a longer effect than PGA2.