Acetyl coenzyme A (acetyl-CoA) carboxylase (ACC) catalyzes carboxylation of acetyl-CoA to form malonyl-CoA. In mammals, two isozymes exist with distinct physiological roles: cytosolic ACC1 participates in de novo lipogenesis (DNL), and mitochondrial ACC2 is involved in negative regulation of mitochondrial β-oxidation. Since systemic ACC1 null mice were embryonic lethal, to clarify the physiological role of ACC1 in hepatic DNL, we generated the liver-specific ACC1 null mouse by crossbreeding of an Acc1^lox(ex46) mouse, in which exon 46 of Acc1 was flanked by two loxP sequences and the liver-specific Cre transgenic mouse. In liver-specific ACC1 null mice, neither hepatic Acc1 mRNA nor protein was detected. However, to compensate for ACC1 function, hepatic ACC2 protein and activity were induced 1.4 and 2.2 times, respectively. Surprisingly, hepatic DNL and malonyl-CoA were maintained at the same physiological levels as in wild-type mice. Furthermore, hepatic DNL was completely inhibited by an ACC1/2 dual inhibitor, 5-tetradecyloxyl-2-furancarboxylic acid. These results strongly demonstrate that malonyl-CoA from ACC2 can access fatty acid synthase and become the substrate for the DNL pathway under the unphysiological circumstances that result with ACC1 disruption. Therefore, there does not appear to be strict compartmentalization of malonyl-CoA from either of the ACC isozymes in the liver.

The functional roles of transient receptor potential (TRP) channels in the gastrointestinal tract have garnered considerable attention in recent years. We previously reported that daikenchuto (TU-100), a traditional Japanese herbal medicine, increased intestinal blood flow (IBF) via adrenomedullin (ADM) release from intestinal epithelial (IE) cells (Kono T et al. J Crohns Colitis 4: 161–170, 2010). TU-100 contains multiple TRP activators. In the present study, therefore, we examined the involvement of TRP channels in the ADM-mediated vasodilatatory effect of TU-100. Rats were treated intraduodenally with the TRP vanilloid type 1 (TRPV1) agonist capsaicin (CAP), the TRP ankyrin 1 (TRPA1) agonist allyl-isothiocyanate (AITC), or TU-100, and jejunum IBF was evaluated using laser-Doppler blood flowmetry. All three compounds resulted in vasodilatation, and the vasodilatory effect of TU-100 was abolished by a TRPA1 antagonist but not by a TRPV1 antagonist. Vasodilatation induced by AITC and TU-100 was abrogated by anti-ADM antibody treatment. RT-PCR and flow cytometry revealed that an IEC-6 cell line originated from the small intestine and purified IE cells expressed ADM and TRPA1 but not TRPV1. AITC increased ADM release in IEC cells remarkably, while CAP had no effect. TU-100 and its ingredient 6-shogaol (6SG) increased ADM release dose-dependently, and the effects were abrogated by a TRPA1 antagonist. 6SG showed similar TRPA1-dependent vasodilatation in vivo. These results indicate that TRPA1 in IE cells may play an important role in controlling bowel microcirculation via ADM release. Epithelial TRPA1 appears to be a promising target for the development of novel strategies for the treatment of various gastrointestinal disorders.

Objective. Chemotherapy-induced oral mucositis (COM) is characterized by painful inflammation with prolonged damage that involves the pathological pain-evoking prostaglandin E2 (PGE2). We previously found that gargling with hangeshashinto (HST), a traditional Japanese medicine, was effective for the treatment of COM. However, little is known regarding the mechanisms. Our aim was to identify the active ingredients and clarify the characteristic effects of HST on the PGE2 system. Methods. Prostanoids produced by human oral keratinocytes (HOK) stimulated with IL-1β were measured by enzyme immunoassay. Active ingredients that regulate PGE2 production were identified and quantified by liquid chromatography–tandem mass spectrometry (LC-MS/MS) and a culture system of HOK cells. Results. Inducible PGE2, PGD2, and PGF2α, metabolites of cyclooxygenase (COX) pathways, were reduced by HST (10-300 µg/mL) without inducing cytotoxicity. The active ingredients of HST were quantified by LC-MS/MS, and [6]-shogaol, [6]-gingerol, wogonin, baicalein, baicalin, and berberine were shown to reduce PGE2 production. A mixture of these 6 ingredients at concentrations equal to 300 µg/mL of HST strongly suppressed PGE2 production to the same level as HST. [6]-Shogaol and [6]-gingerol did not decrease COX-2 mRNA expression and mostly inhibited PGE2 metabolic activity in an assay using intact HOK cells, suggesting that they regulate PGE2 synthesis at the posttranscriptional level. Wogonin, baicalin, and berberine inhibited expression of COX-2 mRNA without affecting PGE2 metabolic activity. Moreover, wogonin, but not [6]-shogaol, suppressed phosphorylation of mitogen-activated protein kinases (p38s and JNKs). Conclusions. These lines show that HST includes several PGE2-regulating ingredients that have different mechanisms and can function as a multicomponent and multitarget agent for treatment of COM, indicating that HST may be beneficial in a new medical strategy for COM treatment.

Various colonic motor activities are thought to mediate propulsion and mixing/absorption of colonic content. The Japanese traditional medicine daikenchuto (TU-100), which is widely used for postoperative ileus in Japan, accelerates colonic emptying in healthy humans. Hydroxy-α sanshool (HAS), a readily absorbable active ingredient of TU-100 and a KCNK3/KCNK9/KCNK18 blocker as well as TRPV1/TRPA1 agonist, has been investigated for its effects on colonic motility. Motility was evaluated by intraluminal pressure and video imaging of rat proximal colons in an organ bath. Distribution of KCNKs was investigated by RT-PCR, in situ hybridization, and immunohistochemistry. Current and membrane potential were evaluated with use of recombinant KCNK3- or KCNK9-expressing Xenopus oocytes and Chinese hamster ovary cells. Defecation frequency in rats was measured. HAS dose dependently induced strong propulsive “squeezing” motility, presumably as long-distance contraction (LDC). TRPV1/TRPA1 agonists induced different motility patterns. The effect of HAS was unaltered by TRPV1/TRPA1 antagonists and desensitization. Lidocaine (a nonselective KCNK blocker) and hydroxy-β sanshool (a geometrical isomer of HAS and KCNK3 blocker) also induced colonic motility as a rhythmic propagating ripple (RPR) and a LDC-like motion, respectively. HAS-induced “LDC,” but not lidocaine-induced “RPR,” was abrogated by a neuroleptic agent tetrodotoxin. KCNK3 and KCNK9 were located mainly in longitudinal smooth muscle cells and in neural cells in the myenteric plexus, respectively. Administration of HAS or TU-100 increased defecation frequency in normal and laparotomy rats. HAS may evoke strong LDC possibly via blockage of the neural KCNK9 channel in the colonic myenteric plexus.

Background: Stimulation of transient receptor potential ankyrin 1 (TRPA1), which abundantly expressed in enterochromaffin cells (ECC), has been reported to exert apparently contradictory results in in vitro contractility and in vivo gastrointestinal (GI) transit evaluations. The pharmaceutical‐grade Japanese traditional medicine daikenchuto (TU‐100) has been reported to be beneficial for postoperative ileus (POI) and accelerate GI transit in animals and humans. TU‐100 was recently shown to increase intestinal blood flow via stimulation of TRPA1 in the epithelial cells of the small intestine (SI).Methods: The effects of various TRPA1 agonists on motility were examined in a manipulation‐induced murine POI model, in vitro culture of SI segments and an ECC model cell line, RIN‐14B.Key Results: Orally administered TRPA1 agonists, aryl isothiocyanate (AITC) and cinnamaldehyde (CA), TU‐100 ingredients, [6]‐shogaol (6S) and γ‐sanshool (GS), improved SI transit in a POI model. The effects of AITC, 6S and GS but not CA were abrogated in TRPA1‐deficient mice. SI segments show periodic peristaltic motor activity whose periodicity disappeared in TRPA1‐deficient mice. TU‐100 augmented the motility. AITC, CA and 6S increased 5‐HT release from isolated SI segments and the effects of all these compounds except for CA were lost in TRPA1‐deficient mice. 6S and GS induced a release of 5‐HT from RIN‐14B cells in a dose‐ and TRPA1‐dependent manner.Conclusions & Inferences: Intraluminal TRPA1 stimulation is a potential therapeutic strategy for GI motility disorders. Further investigation is required to determine whether 5‐HT and/or ECC are involved in the effect of TRPA1 on motility.