Transient receptor potential vanilloid 4 (TRPV4) is a non-selective cation channel that responds to mechanical, thermal, and chemical stimuli in addition to various endogenous ligands, such as arachidonic acid metabolites. The present study aimed to elucidate the expression of TRPV4 in the gastrointestinal tract and the pathogenic roles of TRPV4 in dextran sulphate sodium (DSS)-induced colitis. TRPV4-immunoreactivity was detected in epithelial-like cells of the mouse tongue, esophagus, stomach, ileum, and colon; TRPV4 expression in the tongue was higher than other gastrointestinal tracts. TRPV4 colocalized with a type IV cell marker sonic hedgehog in circumvallate papillae. These findings suggest that TRPV4 contributes to sour taste sensing by regulating type III taste cell differentiation in mice. DSS-induced colitis was significantly attenuated in TRPV4-knockout (TRPV4KO) mice when compared to wild-type mice. DSS treatment upregulated TRPV4 expression in vascular endothelia of colonic mucosa and submucosa. DSS treatment increased vascular permeability, which was abolished in TRPV4KO mice. The activation of TRPV4 decreased VE-cadherin expression in mouse aortic endothelial cells exposed to TNF-α. These findings indicate that the upregulation of TRPV4 in vascular endothelial cells contributes to the progression of colonic inflammation via the activation of vascular permeability. Thus, TRPV4 is an attractive target for the treatment of inflammatory bowel diseases.

Among voltage-gated Ca²⁺ channels, T-type Ca²⁺ channels, which are activated by low voltages, regulate neuronal excitability, spontaneous neurotransmitter release, hormone secretion, etc. and also participate in proliferation of distinct cancer cells. Among three isoforms of T-type Ca²⁺ channels, Ca_v3.2 is detectable in 100% of biopsy samples from prostate cancer patients. In general, prostate cancer cells are highly sensitive to androgen deprivation therapy, but often acquire hormone-therapy resistance. The androgen deprivation may trigger neuroendocrine (NE)-like differentiation of some prostate cancer cells. We have analyzed the expression and function of Ca_v3.2 in human prostate cancer LNCaP cells during NE-like differentiation. NE-like LNCaP cells overexpress Ca_v3.2 through the CREB/Egr-1 pathway and also cystathionine-γ-lyase (CSE), which generates H₂S that enhances the channel activity of Ca_v3.2. H₂S generated by upregulated CSE appears to enhance the activity of upregulated Ca_v3.2 after the differentiation. The enhanced Ca_v3.2 activity in NE-like cells may contribute to increased secretion of mitogenic factors essential for androgen-independent proliferation of surrounding prostate cancer cells. It is known that increased extracellular glucose levels enhance Ca_v3.2 activity through asparagine (N)-linked glycosylation of Ca_v3.2, which might contribute to diabetic neuropathy. We then found that high glucose accelerates the enhanced channel function and overexpression of Ca_v3.2 in NE-like LNCaP cells, which might be associated with clinical evidence for diabetes-related poor prognosis of prostate cancer and development of hormone therapy resistance. Thus, Ca_v3.2 is considered to play a role in the pathophysiology of prostate cancer, and may serve as a therapeutic target.

Digitoxin and digoxin are plant-derived cardiac glycosides. They are Na⁺,K⁺-ATPase (sodium pump) inhibitors, and have been used clinically for treatment and prevention of heart failure and various tachycardia. On the other hand, some epidemiological studies showed that digoxin users have a lower cancer risk compared to the non-users, and that cancer patients who had been treated with digoxin face on improvement of their survival. In various in vitro studies, cardiac glycosides at sub-μM concentrations, which have no significant effect on enzymatic and ion-transporting activities of Na⁺,K⁺-ATPase, show anti-cancer effects. Na⁺,K⁺-ATPase is ubiquitously expressed, so it remains unclear why low concentrations of cardiac glycosides have cancer-specific effects. Recently, we found that the receptor-type Na⁺,K⁺-ATPase, which has no pumping activity, is associated with leucine-rich repeat-containing 8 family, member A(LRRC8A), one of the components of volume-regulated anion channel (VRAC), in the membrane microdomains of plasma membrane of cancer cells, and that this crosstalk contributes to the inhibition of the cancer cell growth by sub-μM cardiac glycosides. In this mechanism, cardiac glycosides bind to the receptor-type Na⁺,K⁺-ATPase, and then stimulate the production of reactive oxygen species (ROS) via NADPH oxidase. The ROS activate VRAC within the membrane microdomains, thus eliciting anti-proliferative effects. VRAC is ubiquitously expressed, and it is normally activated by cell swelling. However, VRAC is activated by cardiac glycoside without cell swelling. On the other hand, the cardiac glycosides-induced effects were not observed in non-cancer cells. Our findings can partly explain why cardiac glycosides elicit selective effects in cancer cells.

Similar to calcium (Ca²⁺) and chloride (Cl⁻) ion channels/transporters, potassium (K⁺) channels have been recognized as a crucial cancer treatment target. Recent studies have provided convincing evidences of positive correlation between elevated expression levels of Ca²⁺-activated K⁺ (K_Ca) channels and cancer proliferation, metastasis, and poor patient prognosis. In cancer cells, K_Ca1.1 and K_Ca3.1 K_Ca channels are co-localized with Ca²⁺-permeable Orai/TRP channels to provide a positive-feedback loop for Ca²⁺ entry. They are responsible for the promotion of cell growth and metastasis in the different types of cancer, and are therefore potential therapeutic targets and biomarkers for cancer. We determined the epigenetic and post-transcriptional dysregulation of K_Ca3.1 by class I histone deacetylase inhibitors in breast and prostate cancer cells. We further determined the transcriptional repression and protein degradation of K_Ca1.1 by vitamin D receptor agonists and androgen receptor antagonists, which are expected as potential therapeutic drugs for triple-negative breast cancer. The anti-inflammatory cytokine, interleukin-10 (IL-10) is an immunosuppressive factor involved in tumorigenesis, and plays a crucial role in escape from tumor immune surveillance. We determined K_Ca3.1 activators are a possible therapeutic option to suppress the tumor-promoting activities of IL-10. These results may provide new insights into cancer treatment focused on Ca²⁺-activated K⁺ channels.

More than twenty years have passed since the demonstration of hydrogen sulfide (H₂S) as a signaling molecule. Various roles of this molecule have been reported including neuromodulation, vascular relaxation, cytoprotection, anti-inflammation, and oxygen sensing. During the study of its effect on neuromodulation, we found TRP channels as a target of H₂S, and later identified polysulfides (H₂S_n) as chemical entity of the ligand. We found that H₂S relaxes vasculatures in synergy with NO, and recently identified H₂S_n as products produced by the chemical interaction between H₂S and NO to exert the effect, suggesting that it may be a mechanism for the synergy between the two molecules. It has attracted attention that sulfite, a further metabolite of H₂S and H₂S_n, protects neurons from oxidative stress by a mechanism different from that by H₂S and H₂S_n.

Hydrogen sulfide (H₂S) has been reported to play an important role in biological systems. More recently, sulfane sulfur (sulfur with 0 or −1 charge) molecules have been also reported to be involved in various biological phenomena such as regulation of redox signaling and antioxidant functions. Fluorescent probes are one of the important chemical tools because it is easy to use and enable the real-time detection of the target molecules in living cells and tissues. We have successfully developed a highly selective H₂S-detecting fluorescent probe, HSip-1. HSip-1 has been designed on the basis of the facts that the macrocyclic polyamine ligands form a stable complex with Cu²⁺, and Cu²⁺ also reacts with H₂S and make a stable CuS complex. SSip-1 is a fluorescent probe for detecting sulfane sulfur and this fluorescent probe is designed on the basis of the unique feature of sulfane sulfur to bind reversibly to other sulfur atoms and the intramolecular spirocyclization reaction of xanthene dyes. SSip-1 is a highly selective fluorescent probe and can detect sulfane sulfur reversibly. Both HSip-1 and SSip-1 were able to be used for the live-cell fluorescence imaging. Further, we applied HSip-1 to the high-throughput screening (HTS) for the inhibitors of 3-mercaptopyruvate sulfurtransferase (3MST), one of the reactive sulfur species (RSS)-generating enzymes. We successfully found new 3MST inhibitors by screening of 174,118 compounds. We expect that these fluorescent probes and inhibitors would be useful to elucidate new functions of RSS and RSS-generating enzymes.

Hydrogen sulfide (H₂S), an endogenous gasotransmitter, is generated from L-cysteine by 3 distinct enzymes including cystathionine-γ-lyase (CSE), and targets multiple molecules, thereby playing various roles in health and disease. H₂S triggers or accelerates somatic pain and visceral nociceptive signals in the pancreas, colon and bladder by enhancing the activity of Ca_v3.2 T-type calcium channels. H₂S also activates TRPA1, which participates in H₂S-induced somatic pain signaling. However, Ca_v3.2 predominantly mediates colonic nociception by H₂S, because genetic deletion of TRPA1 does not reduce H₂S-induced colonic pain. The functional upregulation of the CSE/H₂S/Ca_v3.2 system is involved in neuropathic pain and visceral pain accompanying pancreatitis and cystitis. Ca_v3.2 also appears to participate in irritable bowel syndrome (IBS), although the role of endogenous H₂S generation by CSE in IBS is still open to question. In this review, we describe how H₂S regulates pain signals, particularly by interacting with Ca_v3.2.

Hydrogen sulfide (H₂S) has been focused as a biological mediator, which modulates signal transduction and protects cells and tissues from oxidative stress. H₂S is also expected as a neuroprotectant because it has a neuroprotective activity. Endogenous H₂S is mainly generated from L-cysteine. However, it is difficult to use L-cysteine as a neuroprotectant because of its neurotoxicity. In 2013, a novel biogenesis pathway of H₂S from D-cysteine has been identified. In this pathway, D-amino acid oxidase (DAO) converts D-cysteine to 3-mercaptopyruvate (3MP), followed by the generation of H₂S from 3MP by 3-mercaptopyrvate sulfurtransferase. DAO is especially abundant in cerebellum among various brain regions and mediates efficient generation of H₂S from D-cysteine in the cerebellar tissues. In addition, D-cysteine has more potent neuroprotective activity in cerebellar primary neurons than L-cysteine. Cerebella Purkinje cells (PCs) are characterized by the highly-branched dendrites and are important for cerebellar functions. The dendritic shrinkage and degeneration of PCs are frequently observed in patients and model mice of cerebellar ataxias. We revealed that D-cysteine enhanced dendritic development of primary cultured PCs, but L-cysteine impaired the dendritic development. This effect of D-cysteine was inhibited by DAO inhibitors and reproduced by 3MP and a H₂S donor, suggesting that this enhancement of dendritic development is caused by the production of H₂S from D-cysteine. Taken together, D-cysteine would be available as a neuroprotectant against cerebellar ataxias, which are accompanied with dendritic shrinkage of cerebellar PCs.

Zinc, an essential trace element, plays an important role in a large number of biological functions. In mammalian brain, whereas the majority of brain zinc is bound to proteins including metallothionein, about 5–15% is stored in presynaptic vesicles of glutamatergic neurons throughout the forebrain, especially in the hippocampus, in a relatively free state. Thus, free zinc (Zn²⁺) concentration in the brain is considered to be regulated in order to maintain normal brain functions such as learning and memory. On the other hand, brain Zn²⁺ dyshomeostasis has been recognized as a mechanism for neuronal injury in brain disorders including Alzheimer’s disease and brain ischemia. In particular, after transient brain ischemia, Zn²⁺ accumulates in hippocampal neurons via a zinc transport system, or via release from cytosolic zinc-binding proteins, which results in neuronal cell death. Recently, it has been demonstrated that Zn²⁺ dyshomeostasis also occurs in glial cells such as microglia, astrocytes and oligodendrocytes after brain ischemia. In oligodendrocytes, ischemic insult triggers intracellular Zn²⁺ accumulation, resulting in cell death via mitochondrial dysfunction. Increased extracellular Zn²⁺ inhibits astrocytic glutamate uptake. In addition, extracellular Zn²⁺ massively released from ischemic neurons primes microglia to enhance production of pro-inflammatory cytokines in response to stimuli that trigger M1 activation. This review aims to describe the impact of brain Zn²⁺ dyshomeostasis on alterations in glial cell survival and functions in post-ischemic brains.

Quantitative systems pharmacology (QSP) is an emerging field of modeling technologies that describes the dynamic interaction between biological systems and drugs. Recently, QSP is increasingly being applied to pharmaceutical drug discovery and development, and used for various types of decision makings. In contrast to empirical and statistical models, QSP represents complex systems of human physiology by integrating comprehensive biological information, hence, it can address various purposes including target and/or disease-related biomarker identification, hypothesis testing, and prediction of clinical efficacy or toxicity. On the other hand, structures of QSP models become quite complicated with huge amount of biological components, therefore, close collaboration between pharmacologists having profound knowledge of biology and drug metabolism and pharmacokinetics (DMPK) scientists, experts of model building, is crucial for QSP development and implementation. This article introduces, from DMPK scientists to pharmacologists, main features of QSP and its applications in pharmaceutical industries, and discusses challenges and future perspectives for effective utilization in drug discovery and development.