Establishing Monoclonal Gammopathy of Undetermined Significance as an Independent Pre-Disease State of Multiple Myeloma Using Raman Spectroscopy, Dynamical Network Biomarker Theory, and Energy Landscape Analysis.

Multiple myeloma (MM) is a cancer of plasma cells. Normal (NL) cells are considered to pass through a precancerous state, such as monoclonal gammopathy of undetermined significance (MGUS), before transitioning to MM. In the present study, we acquired Raman spectra at three stages-834 NL, 711 MGUS, and 970 MM spectra-and applied the dynamical network biomarker (DNB) theory to these spectra. The DNB analysis identified MGUS as the unstable pre-disease state of MM and extracted Raman shifts at 1149 and 1527-1530 cm-1 as DNB variables. The distribution of DNB scores for each patient showed a significant difference between the mean values for MGUS and MM patients. Furthermore, an energy landscape (EL) analysis showed that the NL and MM stages were likely to become stable states. Raman spectroscopy, the DNB theory, and, complementarily, the EL analysis will be applicable to the identification of the pre-disease state in clinical samples.

New Possibilities for Evaluating the Development of Age-Related Pathologies Using the Dynamical Network Biomarkers Theory.

Aging is the slowest process in a living organism. During this process, mortality rate increases exponentially due to the accumulation of damage at the cellular level. Cellular senescence is a well-established hallmark of aging, as well as a promising target for preventing aging and age-related diseases. However, mapping the senescent cells in tissues is extremely challenging, as their low abundance, lack of specific markers, and variability arise from heterogeneity. Hence, methodologies for identifying or predicting the development of senescent cells are necessary for achieving healthy aging. A new wave of bioinformatic methodologies based on mathematics/physics theories have been proposed to be applied to aging biology, which is altering the way we approach our understand of aging. Here, we discuss the dynamical network biomarkers (DNB) theory, which allows for the prediction of state transition in complex systems such as living organisms, as well as usage of Raman spectroscopy that offers a non-invasive and label-free imaging, and provide a perspective on potential applications for the study of aging.

Practices, Potential, and Perspectives for Detecting Predisease Using Raman Spectroscopy.

Raman spectroscopy shows great potential for practical clinical applications. By analyzing the structure and composition of molecules through real-time, non-destructive measurements of the scattered light from living cells and tissues, it offers valuable insights. The Raman spectral data directly link to the molecular composition of the cells and tissues and provides a "molecular fingerprint" for various disease states. This review focuses on the practical and clinical applications of Raman spectroscopy, especially in the early detection of human diseases. Identifying predisease, which marks the transition from a healthy to a disease state, is crucial for effective interventions to prevent disease onset. Raman spectroscopy can reveal biological processes occurring during the transition states and may eventually detect the molecular dynamics in predisease conditions.

Allergic inflammation disrupts epithelial electrogenic electrolyte transport through cholinergic regulation in the mouse colon.

Intestinal transport of electrolytes is regulated by the enteric nervous system. Acetylcholine (ACh) is considered the most important neurotransmitter for electrolyte transport in the colon. However, electrolyte transport regulated by ACh is not fully understood in the colon. We investigated the regulation of electrogenic electrolyte transport by cholinergic agonists in the mouse colon by measuring the short-circuit current (Isc) using an Ussing chamber system. Muscarinic stimulation induced transient electrogenic Cl- secretion, and nicotinic stimulation induced electrogenic K+ secretion to the apical side in the normal mouse colon, and these effects were reduced in the colon of mice with food allergy (FA). Administration of prednisolone to mice with FA suppressed mild inflammation in the colon and allergic symptoms and thereby ameliorated the disruption of electrogenic electrolyte transport induced not only by cholinergic pathway activation but also by electrical field stimulation and intracellular cAMP signaling pathway activation in the colon. These results suggest that the electrogenic electrolyte transport function in the colon is impaired by FA-induced colonic inflammation and that the suppression of inflammation ameliorates the dysfunction of electrogenic electrolyte transport in the colon of mice with FA.

Application of the Dynamical Network Biomarker Theory to Raman Spectra.

The dynamical network biomarker (DNB) theory detects the early warning signals of state transitions utilizing fluctuations in and correlations between variables in complex systems. Although the DNB theory has been applied to gene expression in several diseases, destructive testing by microarrays is a critical issue. Therefore, other biological information obtained by non-destructive testing is desirable; one such piece of information is Raman spectra measured by Raman spectroscopy. Raman spectroscopy is a powerful tool in life sciences and many other fields that enable the label-free non-invasive imaging of live cells and tissues along with detailed molecular fingerprints. Naïve and activated T cells have recently been successfully distinguished from each other using Raman spectroscopy without labeling. In the present study, we applied the DNB theory to Raman spectra of T cell activation as a model case. The dataset consisted of Raman spectra of the T cell activation process observed at 0 (naïve T cells), 2, 6, 12, 24 and 48 h (fully activated T cells). In the DNB analysis, the F-test and hierarchical clustering were used to detect the transition state and identify DNB Raman shifts. We successfully detected the transition state at 6 h and related DNB Raman shifts during the T cell activation process. The present results suggest novel applications of the DNB theory to Raman spectra ranging from fundamental research on cellular mechanisms to clinical examinations.

Neuro-immune crosstalk and food allergy: Focus on enteric neurons and mucosal mast cells.

The nervous system and the immune system individually play important roles in regulating the processes necessary to maintain physiological homeostasis, respond to acute stress and protect against external threats. These two regulating systems for maintaining the living body had often been assumed to function independently. Allergies develop as a result of an overreaction of the immune system to substances that are relatively harmless to the body, such as food, pollen and dust mites. Therefore, it has been generally supposed that the development and pathogenesis of allergies can be explained through an immunological interpretation. Recently, however, neuro-immune crosstalk has attracted increasing attention. Consequently, it is becoming clear that there is close morphological proximity and physiological and pathophysiological interactions between neurons and immune cells in various peripheral tissues. Thus, researchers are now beginning to appreciate that neuro-immune interactions may play a role in tissue homeostasis and the pathophysiology of immune-mediated disease, but very little information is available on the molecular basis of these interactions. Mast cells are a part of the innate immune system implicated in allergic reactions and the regulation of host-pathogen interactions. Mast cells are ubiquitous in the body, and these cells are often found in close proximity to nerve fibers in various tissues, including the lamina propria of the intestine. Mast cells and neurons are thought to communicate bidirectionally to modulate neurophysiological effects and mast cell functions, which suggests that neuro-immune interactions may be involved in the pathology of allergic diseases.

Cholinergic anti-inflammatory pathway ameliorates murine experimental Th2-type colitis by suppressing the migration of plasmacytoid dendritic cells.

Ulcerative colitis (UC) is a chronic inflammatory bowel disease. Several studies have demonstrated that α7 nicotinic acetylcholine receptors (α7nAChRs) exert anti-inflammatory effects on immune cells and nicotine suppress UC onset and relapse. Plasmacytoid dendritic cells (pDCs) reportedly accumulate in the colon of UC patients. Therefore, we investigated the pathophysiological roles of α7nAChRs on pDCs in the pathology of UC using oxazolone (OXZ)-induced Th2-type colitis with BALB/c mice. 2-deoxy-D-glucose, a central vagal stimulant suppressed OXZ colitis, and nicotine also ameliorated OXZ colitis with suppressing Th2 cytokines, which was reversed by α7nAChR antagonist methyllycaconitine. Additionally, α7nAChRs were expressed on pDCs, which were located very close to cholinergic nerve fibers in the colon of OXZ mice. Furthermore, nicotine suppressed CCL21-induced bone marrow-derived pDC migration due to Rac 1 inactivation, which was reversed by methyllycaconitine, a JAK2 inhibitor AG490 or caspase-3 inhibitor AZ-10417808. CCL21 was mainly expressed in the isolated lymphoid follicles (ILFs) of the colon during OXZ colitis. The therapeutic effect of cholinergic pathway on OXZ colitis probably through α7nAChRs on pDCs were attributed to the suppression of pDC migration toward the ILFs. Therefore, the activation of α7nAChRs has innovative therapeutic potential for the treatment of UC.

Histopathological Composition of Thrombus in Acute Ischemic Stroke May Vary Even Within the Same Patient: A Preliminary Study Examining Clots According to Their Area of Retrieval.

Objective: Mechanical thrombectomy enables histopathological examination of clots in patients who have suffered acute ischemic strokes. Many studies have described about the relationship between the histopathological compositions of retrieved thrombi and imaging findings, clinical outcomes, and stroke etiology without consensus. In this study, we examined the histological composition of thrombi according to their retrieval site and methods. Methods: We divided retrieved clots into three parts (those retrieved from the proximal and distal parts of the stent retriever, and those aspirated through the guiding catheter) and then histopathologically analyzed their compositions by measuring the area occupied by red blood cells (RBCs), fibrin/platelets (F/Ps), and white blood cells (WBCs). Results: Each specimen showed various composition even within the same patient. For example, the area occupied by RBCs was 20.9% ± 12.1%, 30.5% ± 13.5%, and 41.3% ± 16.1% in the clot retrieved from the proximal and distal parts of the stent retriever, and those aspirated through the guiding catheter, respectively. Conclusion: Histopathological clot composition may vary even within the patient. Further research is needed to investigate more objective methods of histopathological analysis and their clinical significance.

Suppression of plasmacytoid dendritic cell migration to colonic isolated lymphoid follicles abrogates the development of colitis.

BACKGROUND: Dendritic cells (DCs) play a pivotal role in maintaining immunological homeostasis by orchestrating innate and adaptive immune responses via migration to inflamed sites and the lymph nodes (LNs). Plasmacytoid DCs (pDCs) have been reported to accumulate in the colon of inflammatory bowel disease (IBD) patients and dextran sulfate sodium (DSS)-induced colitis mice. However, the role of pDCs in the progression of colonic inflammation remains unclear. METHODS: 80 compounds in natural medicines were searched for inhibitors of pDC migration using bone marrow-derived pDCs (BMpDCs) and conventional DCs (BMcDCs). BALB/c mice were given 3% DSS in the drinking water to induce acute colitis. Compounds, which specifically inhibited pDC migration, were administrated into DSS-induced colitis mice. FINDINGS: Astragaloside IV (As-IV) and oxymatrine (Oxy) suppressed BMpDC migration but not BMcDC migration. In DSS-induced colitis mice, the number of pDCs was markedly increased in the colonic lamina propria (LP), and the expression of CCL21 was obviously observed in colonic isolated lymphoid follicles (ILFs). As-IV and Oxy reduced symptoms of colitis and the accumulation of pDCs in colonic ILFs but not in the colonic LP. Moreover, in a BMpDC adoptive transfer model, BMpDC migration to colonic ILFs was significantly decreased by treatment with As-IV or Oxy. INTERPRETATION: pDCs accumulated in the colon of colitis mice, and As-IV and Oxy ameliorated colitis by suppressing pDC migration to colonic ILFs. Accordingly, the selective inhibition of pDC migration may be a potential therapeutic approach for treating colonic inflammatory diseases.

Pathophysiological Roles of Neuro-Immune Interactions between Enteric Neurons and Mucosal Mast Cells in the Gut of Food Allergy Mice.

Recently, the involvement of the nervous system in the pathology of allergic diseases has attracted increasing interest. However, the precise pathophysiological role of enteric neurons in food allergies has not been elucidated. We report the presence of functional high-affinity IgE receptors (FcεRIs) in enteric neurons. FcεRI immunoreactivities were observed in approximately 70% of cholinergic myenteric neurons from choline acetyltransferase-eGFP mice. Furthermore, stimulation by IgE-antigen elevated intracellular Ca2+ concentration in isolated myenteric neurons from normal mice, suggesting that FcεRIs are capable of activating myenteric neurons. Additionally, the morphological investigation revealed that the majority of mucosal mast cells were in close proximity to enteric nerve fibers in the colonic mucosa of food allergy mice. Next, using a newly developed coculture system of isolated myenteric neurons and mucosal-type bone-marrow-derived mast cells (mBMMCs) with a calcium imaging system, we demonstrated that the stimulation of isolated myenteric neurons by veratridine caused the activation of mBMMCs, which was suppressed by the adenosine A3 receptor antagonist MRE 3008F20. Moreover, the expression of the adenosine A3 receptor gene was detected in mBMMCs. Therefore, in conclusion, it is suggested that, through interaction with mucosal mast cells, IgE-antigen-activated myenteric neurons play a pathological role in further exacerbating the pathology of food allergy.

Corrigendum to "Suppression of Dynamical Network Biomarker Signals at the Predisease State (Mibyou) before Metabolic Syndrome in Mice by a Traditional Japanese Medicine (Kampo Formula) Bofutsushosan".

[This corrects the article DOI: 10.1155/2020/9129134.].

Morphologies and distributions of 5-HT containing enteroendocrine cells in the mouse large intestine.

Serotonin (5-HT)-containing gastrointestinal endocrine cells contribute to regulation of numerous bodily functions, but whether these functions are related to differences in cell shape is not known. The current study identified morphologies and localization of subtypes of 5-HT-containing enteroendocrine cells in the mouse large intestine. 5-HT cells were most frequent in the proximal colon compared with cecum and distal colon. The large intestine harbored both open (O) cells, with apical processes that reached the lumen, and closed (C) cells, not contacting the lumen, classified into O1, O2, and O3 and C1, C2, and C3 cells, by the lengths of their basal processes. O1 and C1 cells, with basal processes sometimes longer that 100 µm, were most common in the distal colon. Their long basal processes ran against the inner surfaces of the mucosal epithelial cells and were strongly immunoreactive for 5-HT; these processes are ideally placed to communicate with the epithelium and to react to mechanical forces. O2 and C2 cells that had similar but shorter basal processes were also most common in the distal colon. O3 and C3 cells had no or very short basal processes. The O3 open type 5-HT cells were abundant in the proximal colon, particularly at the luminal surface, where they could release 5-HT into the lumen to act on luminal 5-HT receptors. Numerous O3 type 5-HT cells occurred in the lower (submucosal) region of the crypts in all segments and might release 5-HT to influence cell renewal in the crypt proliferative zones.

A novel technique for fence-post tube placement in glioma using the robot-guided frameless neuronavigation technique under exoscope surgery: patient series.

BACKGROUND: Robotic technology is increasingly used in neurosurgery. The authors reported a new technique for fence-post tube placement using robot-guided frameless stereotaxic technology with neuronavigation in patients with glioma. OBSERVATIONS: Surgery was performed using the StealthStation S8 linked to the Stealth Autoguide cranial robotic guidance platform and a high-resolution three-dimensional (3D) surgical microscope. A surgical plan was created to determine the removal area using fence-post tube placement at the tumor and normal brain tissue boundary. Using this surgical plan, the robotic system allowed quick and accurate fence-post tube positioning, automatic alignment of the needle insertion and measurement positions in the brain, and quick and accurate puncture needle insertion into the brain tumor. Use of a ventricular drainage tube for the outer needle cylinder allowed placement of the puncture needle in a single operation. Furthermore, use of a high-resolution 3D exoscope allowed the surgeon to simultaneously view the surgical field image and the navigation screen with minimal line-of-sight movement, which improved operative safety. The position memory function of the 3D exoscope allowed easy switching between the exoscope and the microscope and optimal field of view adjustment. LESSONS: Fence-post tube placement using robot-guided frameless stereotaxic technology, neuronavigation, and an exoscope allows precise glioma resection.

Morphological elucidation of short-chain fatty acid receptor GPR41-positive enteric sensory neurons in the colon of mice with dextran sulfate sodium-induced colitis.

Although the etiology of inflammatory bowel disease (IBD) remains unclear, it has generally been accepted that abnormalities in the intestinal immune system and dysbiosis of the gut microbiota are involved in the pathology of IBD. Recently, short-chain fatty acids (SCFAs) produced by gut microbiota were reported to maintain intestinal homeostasis through their receptors, such as GPR41. However, there are contradictory reports about the role of GPR41 in intestinal inflammation. Consequently, the roles of GPR41 in dysbiosis induced by intestinal inflammation remain unclear. Thus, we investigated the distribution of GPR41 in the colonic mucosa of mice with dextran sulfate sodium (DSS)-induced colitis. GPR41-immunoreactive fibrous structures were observed in the colonic lamina propria and muscularis layer of normal mice. In addition, GPR41-immunoreactive fibrous structures partly colocalized with calcitonin gene-related peptide (CGRP; a neurotransmitter of cholinergic enteric sensory neurons)-immunoreactive nerve fibers in the colonic lamina propria, indicating that GPR41 is expressed in cholinergic intrinsic sensory neurons. Furthermore, both GPR41-immunoreactivities and CGRP-immunoreactivities were significantly increased in the lamina propria of the colon in mice with DSS-induced colitis. Interestingly, GPR41-immunoreactivities were often found in close proximity to F4/80+ macrophages in the colonic mucosa of normal mice, and their frequency was elevated in the colonic mucosa of mice with DSS-induced colitis. Therefore, the crosstalk between SCFA-sensing intrinsic sensory neurons and macrophages might be involved in the pathology of acute colitis.

Isoflavones Suppress Cyp26b1 Expression in the Murine Colonic Lamina Propria.

Isoflavones have many biological activities and are major bioactive components of kakkonto, a traditional Japanese herbal medicine. We previously reported that the combined therapy of oral immune therapy (OIT) and kakkonto downregulates the mRNA expression of Cyp26b1, a major retinoic acid (RA)-degrading enzyme, in the colon of food allergy mice and thereby ameliorates allergic symptoms. In this study, we evaluated the effects of various isoflavones on Cyp26b1 expression in primary cultured lamina propria (LP) cells isolated from the mouse colon. The mRNA expression of Cyp26b1 was extremely downregulated by all isoflavones tested in the LP cells except for puerarin. In particular, genistein and genistin markedly suppressed Cyp26b1 mRNA expression without affecting RA-synthesizing enzyme expression. Moreover, to evaluate the effects of isoflavones on allergic reactions, genistein and genistin were administered to ovalbumin (OVA)-induced food allergy mice. Oral administration of genistin suppressed the development of allergic symptoms. These results raise the possibility that isoflavones elevated the level of RA in the colon by inhibiting RA degradation and then the high concentration of RA in the colon might exert immunosuppressive and antiallergic effects on food allergy mice.

Ginger Increases ALDH1A1 Expression and Enhances Retinoic Acid Signaling in a Human Colonic Epithelial Cell Line.

Aldehyde dehydrogenase 1A1 (ALDH1A1) in intestinal epithelial cells (IECs) plays a critical role in regulating immune responses through the production of retinoic acid (RA). However, little is known about its regulation by dietary components. We previously demonstrated that kakkonto, a Japanese traditional herbal medicine, and its constituent puerarin induce the expression of ALDH1A1 mRNA in colonic IECs and thereby attenuate food allergy symptoms in mice. This study aims to investigate the cellular responses of IECs to ALDH1A1 expression as a result of natural food components. The seven medicinal herbs that compose kakkonto were used to treat cultured an IEC line: Caco-2 cells. Expressions levels of ALDH1A1 were analyzed in Caco-2 cells by quantitative RT-PCR, immunocytochemistry and western blotting. Ginger increased the expression levels of ALDH1A1 mRNA and protein in Caco-2 cells. In addition, ginger significantly upregulated the gene expression of retinoic acid receptor (RAR) alpha (RARA), thereby enhancing RA signaling. Furthermore, ginger downregulated the expression of histone deacetylase (HDAC)2 (HDAC2) and HDAC3 in Caco-2 cells. The present study suggests the possibility that food ingredients such as a ginger modulate vitamin A metabolism in the gut through the regulation of RA synthesis, which may contribute to RA-mediated regulation of immune responses and the regulation of allergic inflammation.

Cisplatin-related Cerebral Infarction in Carcinoma of the External Auditory Canal.

BACKGROUND: Despite the known association between cisplatin and vascular toxicity, the mechanism of cisplatin-associated cerebral infarction, a relatively rare complication, remains unclear. We describe an investigation of potential biomarkers that could facilitate the early detection of this complication in a relevant case. CASE DESCRIPTION: A 59-year-old male diagnosed with stage III carcinoma of the external auditory canal underwent cisplatin chemotherapy. Seven days after the last dose, he presented with a disturbance of consciousness due to basilar artery occlusion, which was associated with chemotherapy administration. The patient recovered consciousness after thrombectomy. Interestingly, an increase in serum von Willebrand factor (vWf) activity was observed. The vWf activity level gradually normalized 5 months after cisplatin administration. CONCLUSIONS: Endothelial injuries could be responsible for cisplatin-associated cerebral infarction. Moreover, a cisplatin-induced cerebral infarction increase in serum vWf activity, which indicates endothelial injury, suggests that this molecule might be a useful biomarker for predicting cisplatin-associated cerebral infarction.

Therapeutic Benefit in Allergic Dermatitis Derived from the Inhibitory Effect of Byakkokaninjinto on the Migration of Plasmacytoid Dendritic Cells.

Dendritic cells (DCs) are well known to be essential immunocytes involved in innate and adaptive immunity. DCs are classified as conventional dendritic cells (cDCs) and plasmacytoid dendritic cells (pDCs). Recently, the accumulation of pDCs in inflamed tissues and lymphoid tissues has been considered to be a possible contributing factor in the development of immunological diseases, but little is known about the pathophysiological roles of pDCs in immunological diseases. To date, many studies have demonstrated that many kinds of Kampo formulas can regulate immunological reactions in human immune diseases. Thus, we screened Kampo formulas to identify an agent that inhibits pDC migration. Furthermore, we investigated the therapeutic effects of these formulas on a murine DNFB-induced allergic contact dermatitis model. Bone marrow-derived pDCs (BMpDCs) were derived from the bone marrow cells of BALB/c mice in a culture medium with Flt3 ligand. The effects of Kampo formulas on BMpDC migration were evaluated by assessing the number, velocity, and directionality of BMpDCs chemotaxing toward the more concentrated side of a chemokine (C-C motif) ligand 21 (CCL21) gradient. The Kampo formulas that exerted inhibitory effects on pDC migration were orally administered to DNFB-induced allergic contact dermatitis model mice. Byakkokaninjinto reduced the number of migrated BMpDCs and suppressed the velocity and directionality of BMpDC migration in a chemotaxis assay. Gypsum Fibrosum and Ginseng Radix, which are components of byakkokaninjinto, obviously suppressed the velocity of BMpDC migration. Furthermore, Gypsum Fibrosum significantly suppressed the directionality of BMpDC migration. In DNFB-induced allergic contact dermatitis model mice, byakkokaninjinto markedly abrogated ear swelling in late-phase allergic reactions. In conclusions, byakkokaninjinto, which has an inhibitory effect on pDC migration, was able to prevent the occurrence of allergic contact dermatitis, suggesting that pDCs were involved in the onset of allergic contact dermatitis in the mouse model. Therefore, byakkokaninjinto is anticipated to be a therapeutic agent for disorders related to pDC migration.

Interleukin-4 Receptor α Subunit Deficiency Alleviates Murine Intestinal Inflammation In Vivo Through the Enhancement of Intestinal Mucosal Barrier Function.

Disturbance of epithelial barrier function causes chronic intestinal inflammation such as inflammatory bowel disease. Several studies have reported that Th2 cytokines such as interleukin (IL)-4 and IL-13 play an important role in the regulation of intestinal barrier function. However, the precise role of the IL-4 receptor α subunit (IL-4Rα) in intestinal inflammation remains unclear. Thus, we used an experimental colitis model to investigate the role of IL-4Rα in intestinal inflammation. IL-4Rα-deficient (IL-4Rα-/-) mice and their littermate wild-type (WT) mice were used. Experimental colitis was induced by administration of 3% dextran sulfate sodium (DSS) in the drinking water for seven days. Treatment with DSS caused body weight loss, an increase in the disease activity index and histological abnormalities in WT colitis mice, all of which were significantly attenuated in IL-4Rα-/- colitis mice. Neutrophil infiltration in the colonic mucosa was reduced in IL-4Rα-/- colitis mice compared with WT colitis mice. NADPH oxidase 1 expression and reactive oxygen species production were increased in the colons of IL-4Rα-/- mice. Furthermore, elevated intestinal permeability induced by DSS treatment was suppressed in IL-4Rα-/- colitis mice. These results demonstrate that IL-4Rα-/- mice exhibit reduced susceptibility to DSS-induced colitis. Our present findings suggest that IL-4Rα deficiency enhances intestinal mucosal barrier function through the upregulation of NADPH oxidase 1-dependent reactive oxygen species production, thereby suppressing the development of intestinal inflammation.

Suppression of Dynamical Network Biomarker Signals at the Predisease State (Mibyou) before Metabolic Syndrome in Mice by a Traditional Japanese Medicine (Kampo Formula) Bofutsushosan.

Due to the increasing incidence of metabolic syndrome, the development of new therapeutic strategies is urgently required. One promising approach is to focus on the predisease state (so-called Mibyou in traditional Japanese medicine) before metabolic syndrome as a preemptive medical target. We recently succeeded in detecting a predisease state before metabolic syndrome using a mathematical theory called the dynamical network biomarker (DNB) theory. The detected predisease state was characterized by 147 DNB genes among a total of 24,217 genes in TSOD (Tsumura-Suzuki Obese Diabetes) mice, a well-accepted model of metabolic syndrome, at 5 weeks of age. The timing of the predisease state was much earlier than the onset of metabolic syndrome in TSOD mice reported to be at approximately 8-12 weeks of age. In the present study, we investigated whether the predisease state in TSOD mice can be inhibited by the oral administration of a Kampo formula, bofutsushosan (BTS), which is usually used to treat obese patients with metabolic syndrome in Japan, from 3 to 7 weeks of age. We found the comprehensive suppression of the early warning signals of the DNB genes by BTS at 5 weeks of age and later. Specifically, the standard deviations of 134 genes among the 147 DNB genes decreased at 5 weeks of age as compared to the nontreatment control group, and 80 of them showed more than 50% reduction. In addition, at 7 weeks of age, the body weight and blood glucose level were significantly lower in the BTS-treated group than in the nontreatment control group. The results of our study suggest a novel mechanism of BTS; it suppressed fluctuations of the DNB genes at the predisease state before metabolic syndrome and thus prevented the subsequent transition to metabolic syndrome. In conclusion, this study demonstrated the preventive and preemptive effects of a Kampo formula on Mibyou before metabolic syndrome for the first time based on scientific evaluation.