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 chromatographytandem 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.

‘Oketsu’ is a pathophysiologic concept in Japanese traditional (Kampo) medicine, primarily denoting blood stasis/stagnant syndrome. Here we have explored plasma protein biomarkers and/or diagnostic algorithms for ‘Oketsu’. Sixteen rheumatoid arthritis (RA) patients were treated with keishibukuryogan (KBG), a representative Kampo medicine for improving ‘Oketsu’. Plasma samples were diagnosed as either having an ‘Oketsu’ (n = 19) or ‘non-Oketsu’ (n = 29) state according to Terasawa's ‘Oketsu’ scoring system. Protein profiles were obtained by surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF MS) and hierarchical clustering and decision tree analyses were performed. KBG treatment for 4 or 12 weeks decreased the ‘Oketsu’ scores significantly. SELDI protein profiles gave 266 protein peaks, whose expression was significantly different between the ‘Oketsu’ and ‘non-Oketsu’ states. Hierarchical clustering gave three major clusters (I, II, III). The majority (68.4%) of ‘Oketsu’ samples were clustered into one cluster as the principal component of cluster I. The remaining ‘Oketsu’ profiles constituted a minor component of cluster II and were all derived from patients cured of the ‘Oketsu’ state at 12 weeks. Construction of the decision tree addressed the possibility of developing a diagnostic algorithm for ‘Oketsu’. A reduction in measurement/pre-processing conditions (from 55 to 16) gave a similar outcome in the clustering and decision tree analyses. The present study suggests that the pathophysiologic concept of Kampo medicine ‘Oketsu’ has a physical basis in terms of the profile of blood proteins. It may be possible to establish a set of objective criteria for diagnosing ‘Oketsu’ using a combination of proteomic and bioinformatics-based classification methods.

Oral mucositis (OM) in cancer patients induced by chemotherapy or radiotherapy has a significant impact on quality of life, and causes considerable morbidity. Oral microorganisms are likely to intensify the inflammatory process and aggravate the formation of ulcers. Hangeshashinto (HST), a Japanese kampo medicine, has been reported to be effective when used as a gargle for the treatment of OM. To clarify the effects of HST on oral microorganisms, we assessed its antimicrobial activity against 27 microbial species, including 19 oral bacteria and one fungus. HST extract inhibited the growth of Gram-negative bacteria, including Fusobacterium nucleatum, Porphyromonas gingivalis, Porphyromonas endodontalis, Prevotella intermedia, Prevotella melaninogenica, Tannerella forsythia, Treponema denticola, and Porphyromonas asaccharolytica, though inhibitory effects were less pronounced for Gram-positive bacteria and the fungal strain. We then investigated the effects of antibacterial activities on 15 purified ingredients of HST and determined that baicalein, berberine, coptisine, [6]-shogaol, and homogentisic acid actively inhibited the growth of these bacteria. These findings showed that HST inhibits the growth of specific Gram-negative periodontopathogenic bacteria, which are significant pathogens in OM, without disturbing the normal oral flora. Our data suggest that HST may be a useful treatment for OM in patients undergoing anticancer treatment.

Oral mucositis (OM) is a common and painful complication of radiotherapy for head and neck cancer. Hangeshashinto (HST), a Japanese traditional medicine, is known to alleviate radiotherapy- and/or chemotherapy-induced OM; however, the detailed mechanism has not yet been clarified. The aim of the present study was to clarify the details of the antioxidative functions of HST against reactive oxygen species (ROS) produced by radiation. The hydroxyl radical (•OH)–scavenging ability and the reduction ability was simultaneously measured using a modified electron paramagnetic resonance (EPR) spin-trapping method. The superoxide (O2•−)–scavenging ability was estimated by an EPR redox probing method. Water suspensions of powdered HST and of its seven constitutive crude drugs were tested. In addition, some of the main water-soluble ingredients of the crude drugs were also tested. HST was found to scavenge both •OH and O2•−. Furthermore, HST was observed to reduce relatively stable nitroxyl radicals. Glycyrrhizae Radix (kanzo), Ginseng Radix (ninjin), Zizyphi Fructus (taiso) and glycyrrhizin (an ingredient of kanzo) were all found to be relatively good •OH scavengers. Scutellariae Radix (ogon) and Coptidis Rhizoma (oren) demonstrated reducing ability. In addition, acteoside and berberine chloride, which are water-soluble ingredients of ogon and oren, respectively, also demonstrated reducing ability. Oren exhibited oxidative ability at higher concentrations, which may have a function in maintaining catalytic redox action. The antioxidative function of HST probably worked via a balance of scavenging ROS, reducing stable free radicals, and some minor oxidizing activities.

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