Procyanidins and sensory nutrition; do procyanidins modulate homeostasis via astringent taste receptors?

Long-term intake of procyanidins has been suggested to reduce the risk of cardiovascular disease, dementia, and sensory function decline associated with aging. However, most of the ingested procyanidins are not absorbed and are excreted in the feces, so the mechanism of their beneficial impact is unknown. Procyanidins are the components of astringency in plant foods and their stimulation appears to be directly transmitted to the central nervous system via sensory nerves. Recent attention has been focused on the taste receptors expressed in the extra-oral gastrointestinal tract may regulate homeostasis via the neuroendocrine system. In this paper, we have reviewed recent findings on the relationship between the astringency of procyanidins and their bioregulatory effects.

Impact of cyanidin 3-O-glucoside on rat micro-and systemic circulation, possibly thorough angiogenesis.

Cyanidin 3-O-glucoside (C3G), an antioxidant, is one of the most abundant anthocyanin in plant foods. Intervention trials and subsequent meta-analyses have suggested that anthocyanins could reduce the risks of cardiovascular diseases. This study investigated hemodynamic alterations following a single intragastric dose of C3G by measuring blood flow in rat cremaster muscle arteriole for 60 min. Next, in excised aortas, we performed western blotting to measure the phosphorylation of Akt and endothelial nitric oxide synthase (eNOS). A single oral dose of C3G significantly increased blood flow soon after ingestion, and it was maintained throughout the experimental period. In addition, aortic Akt phosphorylation increased. Then, we examined the impact of repeated oral administrations of C3G for 14 days. The mean blood pressure was significantly reduced at 7 and 14 days after treatment, with a slight increase in aortic eNOS expression. Immunohistochemical analyses of the soleus showed that the level of CD31, an angiogenesis-marker protein, was significantly increased with C3G. These results suggested that an oral dose of C3G increased blood flow, which promoted angiogenesis within skeletal muscle, and consequently, blood pressure was reduced.

Cinnamtannin A2, (-)-epicatechin tetramer, attenuates skeletal muscle wasting in disuse atrophy model mice induced by hindlimb suspension.

The impact of repeated administration of cinntamtannin A2 (A2, 25 µg/kg) on skeletal muscle disuse atrophy model mice induced by hindlimb suspension for 14 days was examined. In soleus, weight loss and a reduction in the average myofibre size with shifting to the smaller side of the peak were observed in the suspension-vehicle group, but A2 reduced these changes. Average myofibre size significantly increased in ground-A2 compared to ground-vehicle. A marked increase in the dephosphorylation of forkhead box O (FoxO) 3a by the suspension was reduced by A2. The phosphorylation of protein kinase B (Akt) and eukaryotic translation initiation factor 4E-binding protein (4EBP)-1 were significantly increased by the treatment of A2. In addition, a single dose of A2 increased dramatically in the 24-h excretion of catecholamines in urine. These results suggest that A2 administration results in sympathetic nerve activation and promotes hypertrophy while inhibiting the progress of disuse muscle atrophy.

Theaflavins decrease skeletal muscle wasting in disuse atrophy induced by hindlimb suspension in mice.

We previously found that a single dose of theaflavins induced skeletal muscle metabolic changes. In this study, we examined the effect of theaflavins on disuse muscle atrophy model mice by hindlimb suspension. Mice were assigned to 4 groups; ground-vehicle, ground-theaflavins, suspension-vehicle, and suspension-theaflavins, dosed with theaflavins (250 mg/kg/day) for 2 weeks. The peak of myotube size of cross sectional area was significantly moved to the smaller side in the suspension-vehicle group compared with the ground-vehicle group, and these shifts were significantly reduced by the treatment with theaflavins in both soleus and extensor digitorum longus. The level of phosphorylated eukaryotic translation initiation factor 4E-binding protein (4EBP)-1, located downstream of the Akt/mTOR pathway, was significantly different between suspension-vehicle and suspension-theaflavins in soleus. The ratio of forkhead box O (FoxO) 3a to phosphorylated FoxO3a significantly increased in soleus or tended to rise in extensor digitorum longus of suspension-vehicle group compared with ground-vehicle. In contrast, these changes were not observed in suspension-theaflavins group. These results suggested that theaflavins inhibited the progress of disuse muscle atrophy through modulation of protein metabolism.

Sensory Nutrition and Bitterness and Astringency of Polyphenols.

Recent studies have demonstrated that the interaction of dietary constituents with taste and olfactory receptors and nociceptors expressed in the oral cavity, nasal cavity and gastrointestinal tract regulate homeostasis through activation of the neuroendocrine system. Polyphenols, of which 8000 have been identified to date, represent the greatest diversity of secondary metabolites in plants, most of which are bitter and some of them astringent. Epidemiological studies have shown that polyphenol intake contributes to maintaining and improving cardiovascular, cognitive and sensory health. However, because polyphenols have very low bioavailability, the mechanisms of their beneficial effects are unknown. In this review, we focused on the taste of polyphenols from the perspective of sensory nutrition, summarized the results of previous studies on their relationship with bioregulation and discussed their future potential.

Difference in endocrine and behavior between short-term single- and paired-housing mice in metabolic cage.

Metabolic cage housing which is exposed to a number of environmental stressors is often used in pharmacokinetic studies. In this study, we compared the difference in stress response between single- and paired-housing in metabolic cages by evaluating the alteration of urinary stress hormones and behavior. Mice were randomly divided into single- or paired-housing groups and placed in a metabolic cage with wire mesh. Their urine was collected every 24 h for consecutive 4 days to determine excreted catecholamine and corticosterone. The change in body weight was significantly decreased at 3 and 4 days in the single-housing group compared with that before the experiment, but not paired-housing group. The level of urinary catecholamines, such as noradrenaline, adrenaline, and their metabolite vanillylmandelic acid, was significantly increased in the single-housing compared with paired housing group and urinary corticosterone increased as well. Next, for the two similarly housed groups, we observed spontaneous behavior on the fourth day and conducted an elevated plus-maze test on the fifth day. Spontaneous behavior was not different between experimental groups. In the elevated plus-maze test, the proportion of time spent in the open arms was significantly prolonged in the paired-housing group compared to that of the single-housing group. Short-term social isolation stress loading in metabolic cages was suggested to exhibit endocrinological and behavioral changes in mice. To reduce such interference due to stress exposure, it was suggested to keep two mice in a metabolic cage.

Activation of transient receptor potential channels is involved in reactive oxygen species (ROS)-dependent regulation of blood flow by (-)-epicatechin tetramer cinnamtannin A2.

Intervention trials confirmed that blood flow-mediated dilatation increases significantly after intake of astringent (-)-epicatechin (EC) oligomers (procyanidins)-rich foods, but the mechanism remains unclear. We have previously found that procyanidins can activate the sympathetic nervous and subsequently increase blood flow. Here, we examined whether procyanidin-derived reactive oxygen species (ROS) activate transient receptor potential (TRP) channels in gastrointestinal sensory nerves and consequently induce sympathoexcitation. We evaluated the redox properties of EC and its tetramer cinntamtannin A2 (A2) at pH 5 or 7, mimicking plant vacuole or oral cavity/small intestine using a luminescent probe. At pH 5, A2 or EC showed O2·- scavenging ability, but they promoted O2·- generation at pH 7. We observed blood flow in rat cremaster arterioles using laser Doppler, a single oral dose of 10 µg/kg A2 markedly increased blood flow, while EC showed little activity. This change with A2 was significantly dampened by co-administration of adrenaline blocker, ROS scavenger N-acetyl-L-cysteine (NAC), TRP vanilloid 1, or ankyrin 1 antagonist. We also performed a docking simulation of EC or A2 with the binding site of a typical ligand for each TRP channel and calculated the respective binding affinities. The binding energies were notably higher for A2 than typical ligands, suggesting that A2 is less likely to bind to these sites. ROS produced at neutral pH following the orally administered A2 to the gastrointestinal tract could activate TRP channels, triggering sympathetic hyperactivation and causing hemodynamic changes.

Exploring Novel Vitamin K Derivatives with Anti-SARS-CoV-2 Activity.

From our compound library of vitamin K derivatives, we found that some compounds exhibited anti-SARS-CoV-2 activity in VeroE6/TMPRSS2 cells. The common structure of these compounds was menaquinone-2 (MK-2) with either the m-methylphenyl or the 1-naphthyl group introduced at the end of the side chain. Therefore, new vitamin K derivatives having more potent anti-SARS-CoV-2 activity were explored by introducing various functional groups at the ω-position of the side chain. MK-2 derivatives with a purine moiety showed the most potent antiviral activity among the derivatives. We also found that their mechanism of action was the inhibition of RNA-dependent RNA polymerase (RdRp) of SARS-CoV-2. The chemical structures of our compounds were completely different from those of nucleic acid derivatives such as remdesivir and molnupiravir, clinically approved RdRp inhibitors for COVID-19 treatment, suggesting that our compounds may be effective against viruses resistant to these nucleic acid derivatives.

Hormetic response to B-type procyanidin ingestion involves stress-related neuromodulation via the gut-brain axis: Preclinical and clinical observations.

B-type procyanidins, a series of catechin oligomers, are among the most ingested polyphenols in the human diet. Results of meta-analyses have suggested that intake of B-type procyanidins reduces cardiovascular disease risk. Another recent focus has been on the effects of B-type procyanidins on central nervous system (CNS) function. Although long-term B-type procyanidin ingestion is linked to health benefits, a single oral intake has been reported to cause physiological alterations in circulation, metabolism, and the CNS. Comprehensive analyses of previous reports indicate an optimal mid-range dose for the hemodynamic effects of B-type procyanidins, with null responses at lower or higher doses, suggesting hormesis. Indeed, polyphenols, including B-type procyanidins, elicit hormetic responses in vitro, but animal and clinical studies are limited. Hormesis of hemodynamic and metabolic responses to B-type procyanidins was recently confirmed in animal studies, however, and our work has linked these effects to the CNS. Here, we evaluate the hormetic response elicited by B-type procyanidins, recontextualizing the results of intervention trials. In addition, we discuss the possibility that this hormetic response to B-type procyanidins arises via CNS neurotransmitter receptors. We have verified the direction of future research for B-type procyanidins in this review.

Relationship between hemodynamic alteration and sympathetic nerve activation following a single oral dose of cinnamtannin A2.

We previously found that a single dose of B-type procyanidin mixture increase in skeletal muscle blood flow (BF). We compared BF changes following administration of (-)-epicatechin (EC, monomer) and the B-type procyanidins procyanidin B2 (B2, dimer), procyanidin C1 (C1, trimer), and cinnamtannin A2 (A2, tetramer). Each chemical was administered orally to rats, followed by BF measurement in cremaster arteriole for 60 min. About 10 and 100 µg/kg of B2 and C1 elicited BF increase, the effect was potent at 100 µg/kg. BF also increased significantly after administration of 10 µg/kg A2, but not with the administration at 100 µg/kg. EC yielded no BF changes. Co-treatment with the nonselective adrenaline blocker carvedilol attenuated the BF increase seen with 10 µg/kg A2 treatment. This outcome suggested the involvement of sympathetic nerve activation in the BF increase by this dose of A2. Co-treatment of 100 µg/kg A2 with the α2 blocker yohimbine exhibited an increase of BF significantly. The α2 adrenaline receptor in the vasomotor centre is an inhibitory receptor and it regulates hemodynamics. This result suggested that high doses of A2 did not alter BF because of activating the α2 adrenergic receptor. Phosphorylation of aortic endothelial nitric oxide synthase (eNOS) increased with 10 µg/kg A2 alone or co-treatment with 100 µg/kg A2 and yohimbine, but not with co-treatment of 10 µg/kg A2 and carvedilol or 100 µg/kg A2 alone. These results imply that A2 does not directly activate eNOS, but that shear stress from the increased BF might be associated with eNOS phosphorylation.

Repeated Oral Administration of Flavan-3-ols Induces Browning in Mice Adipose Tissues through Sympathetic Nerve Activation.

We previously found increases in uncoupling protein (Ucp)-1 transcription in brown adipose tissue (BAT) of mice following a single oral dose of flavan 3-ol (FL)s, a fraction of catechins and procyanidins. It was confirmed that these changes were totally reduced by co-treatment of adrenaline blockers. According to these previous results, FLs possibly activate sympathetic nervous system (SNS). In this study, we confirmed the marked increase in urinary catecholamine (CA) s projecting SNS activity following a single dose of 50 mg/kg FLs. In addition, we examined the impact of the repeated administration of 50 mg/kg FLs for 14 days on adipose tissues in mice. In BAT, FLs tended to increase the level of Ucp-1 along with significant increase of thermogenic transcriptome factors expressions, such as peroxisome proliferator-activated receptor γ coactivator (PGC)-1α and PR domain-containing (PRDM)1. Expression of browning markers, CD137 and transmembrane protein (TMEM) 26, in addition to PGC-1α were increased in epididymal adipose (eWAT) by FLs. A multilocular morphology with cell size reduction was shown in the inguinal adipose (iWAT), together with increasing the level of Ucp-1 by FLs. These results exert that FLs induce browning in adipose, and this change is possibly produced by the activation of the SNS.

Method for detecting hemodynamic alterations following a single gavage in rats.

It is known that administering a gavage to rodents evokes a cardiac reflex, due to gastrointestinal stimulation. Consequently, it is difficult to evaluate changes in hemodynamics after a single oral dose of a pungent or astringent, which alters the circulation by increasing sympathetic activity. In the present study, we developed a method for administering a gavage without significantly affecting hemodynamics measurements. We marked a gastric tube at 10 cm from the tip, to mark the distance from the oral cavity to the stomach body of Wistar male rats. Rats were intubated under urethane anesthesia.After 10-15 min of stabilization, we measured the mean blood pressure (MBP), heart rate (HR), and blood flow (BF) in the cremaster arteriole under two different conditions; condition 1: a pointed gastric tube, room temperature distilled water, and injected at normal speed (approximately 3 ml/min); condition 2: a rounded gastric tube, 37°C distilled water, and injection at 1.0 ml/min. Under condition 1, we observed striking hemodynamic alterations, due to the somatic afferent reflex. In contrast, under condition 2, these hemodynamic changes were nearly eliminated. In addition, we could clearly detect hemodynamic changes in rats after a single gavage treatment of pungent (capsaicin) or astringent (cinnamtannin A2). We observed transient increases in the HR and MBP soon after treatment with capsaicin. Moreover, cremasteric BF was elevated with cinnamtannin A2. These results confirmed the utility of the gavage method developed in this study.

Acylated anthocyanins derived from purple carrot (Daucus carota L.) induce elevation of blood flow in rat cremaster arteriole.

Acylated anthocyanins are more stable than monomeric anthocyanins, but little is known about their physiological effects. We evaluated the hemodynamic effects of single intragastric doses of purple carrot (Daucus carota L.) anthocyanin (PCA) and two monomeric anthocyanins, cyanidin 3-O-glycoside (C3G) and delphinidin 3-O-ruthenoside (D3R). PCA, C3G, or D3R was administered orally to rat and blood flow in the cremaster artery was measured for 60 min using a laser Doppler blood flow meter. After measurements, the aorta of the animal was removed and the extent of phosphorylation of aortic epithelial nitric oxide synthase (eNOS) and Akt were determined by western blotting. PCA (10 mg kg-1) or C3G (1 mg kg-1) significantly increased rat cremaster arteriole blood flow and phosphorylation of eNOS and Akt; D3G (1 mg kg-1) only slightly altered cremaster arteriole blood flow and did not affect the phosphorylation of eNOS and Akt in the aorta. These results suggest that hemodynamic alterations depend more on the chemical structure of anthocyanins, particularly the aglycon, than on the glycoside. In addition, increase of blood flow by a single oral dose of PCA was practically reduced with treatment of carvedilol (CR), a non-specific adrenaline blocker. Blood concentrations of cyanidin or its glycoside 15, 30, or 60 min after the administration of 10 mg kg-1 PCA were below the limit of detection. These hemodynamic changes may have been associated with an indirect adrenergic action induced following a single dose of PCA.