Oxidative stress has been reported to be associated with the progression of renal pathology as well as with the onset of complications associated with this condition. Bardoxolone methyl, a nuclear factor-erythroid 2-related factor 2 (Nrf2) activator with anti-oxidative and inflammatory modulation effects, has been reported to improve renal function in clinical trials. As of this writing, there have been no systems for the quantitative evaluation of oxidative stress that could be applied as a clinical test. We recently reported on post-translational modifications of albumin using electrospray-ionization time-of-flight mass spectrometry (ESI-TOF MS) and the results indicated that oxidized albumin (cysteinylated albumin: a molecule that was oxidatively cysteinylated at Cys34) was found to increase with the progression of renal pathology. A clinical study demonstrated that monitoring the levels of oxidized albumin could be useful for the early diagnosis of diabetic kidney disease and for predicting renal prognosis. Higher levels of serum oxidized albumin were also associated with cardiovascular complications or sarcopenia in patients who were undergoing hemodialysis. The overall findings indicate that oxidized albumin could also be an index of therapeutic efficacy against kidney disease. Oxidized albumin is not only a marker for kidney disease but also a nephrotoxic substance. This suggests that measuring the levels of oxidized albumin could be a biomarker for diagnosing the progression of and complications associated with renal disease in addition to evaluating the efficacy of therapeutic drugs.

Osteosarcoma (OS) is the most common primary malignant bone tumor and is mainly diagnosed in children. Toll-like receptor 9 (TLR9) is expressed in various tumor cells and was correlated with cancer progression. However, the underlying mechanism of TLR9 on the OS progression remains unclear. Our previous study demonstrated that the expression of TLR9 was positively correlated with the development stage of OS. Herein, we further evaluated the actual roles and the molecular mechanism of TLR9 on regulating OS cell proliferation and metastasis. Our data showed that TLR9 was upregulated in OS cells compared to normal osteoblastic cells, and knockdown of TLR9 inhibited OS cell proliferation and induced cell cycle arrest by the decreased expression of cyclin D1, CDK2, and p-Rb, while TLR9 overexpression exerted the inverse effects. Furthermore, TLR9 overexpression could enhance the migration and invasion activities of the OS cells by the upregulation of matrix metalloproteinases 2 (MMP2) and MMP9, and the opposite result was observed in TLR9-silenced cells. Moreover, the nuclear factor kappa B (NF-κB) signaling pathway was activated by TLR9, and TLR9-induced malignant phenotype of OS cells was abrogated by the NF-κB antagonist BAY11-7082. Our study indicated that TLR9 might play a critical role in facilitating OS progression by activating the NF-κB signaling pathway, which may provide a valuable therapeutic target for OS.

Delavatine A (DA) is an unusual isoquinoline alkaloid with a novel skeleton isolated from Chinese folk medicine Incarvillea delavayi. Studies conducted in our lab have demonstrated that DA has potential anti-inflammatory activity in lipopolysaccharide (LPS)-treated BV-2 cells. DA, however, has not been studied for its protective effect on neuronal cells yet. Thus, to explore whether DA can protect neurons, oxygen and glucose deprivation/reperfusion (OGD/R)-injured PC12 cell and middle cerebral artery occlusion/reperfusion (MCAO/R) rat model were used to assess the protective efficacy of DA against OGD/R damaged PC12 cells and MCAO/R injured rats. Our results demonstrated that DA pretreatment (0.31–2.5 µM) dose-dependently increased cell survival and mitochondrial membrane potential (MMP), whereas it lowered the leakage of lactate dehydrogenase (LDH), intracellular cumulation of Ca²⁺, and overproduction of reactive oxygen species (ROS), and inhibited the apoptosis rate in OGD/R-injured PC12 cells. Western blot demonstrated that DA pretreatment lowered the expression of apoptotic proteins and repressed the activation of the mitogen-activated protein kinase kinase 7 (MKK7)/c-Jun N-terminal kinase (JNK) pathway. It was also found that the neuroprotective efficacy of DA was significantly reversed by co-treatment with the JNK agonist anisomycin, suggesting that DA reduced PC12 cell injury and apoptosis by suppressing the MKK7/JNK pathway. Furthermore, DA oral administration greatly alleviated the neurological dysfunction and reduced the infarct volume of MCAO/R rats. Taken together, DA could ameliorate OGD/R-caused PC12 cell injury and improve brain ischemia/reperfusion (I/R) damage in MCAO/R rats, and its neuroprotection might be attributed to suppressing the MKK7/JNK signaling pathway.

Pentazocine (PTZ) is a widely used drug for postoperative pain. It should be administered at appropriate dosing intervals not only because of its morphine-like side effects but also because frequent inappropriate dosing can lead to dependence. Although perioperative patients reportedly have nonnegligible effects on placebo drugs and postoperative wound healing, no pharmacokinetic (PK)/pharmacodynamic (PD) model has been established and simulated using real-world data for the perioperative period. This study aimed to perform PTZ modeling and simulation and to establish an indicator of the timing of drug efficacy evaluation in clinical practice. Participants were in-hospital orthopedic surgery patients who received 15 mg of PTZ within 48 h postoperatively. Pain severity was assessed using the numerical rating scale (NRS). A two-compartment model was selected for the population PK model and an indirect response model for the PK/PD model. Using these models, a virtual population of 1000 patients with Pain_base NRS of 5 and 6 and body weights of 40, 80, and 120 kg were treated with single and multiple PTZ administrations (4, 8, and 24 h apart) of 15 mg. Simulation results indicate that its analgesic efficacy should be evaluated within 1 h after administration of 15 mg of PTZ. Additional doses should be considered every 8–12 h in postoperative patients with Pain_base NRS of 5 weighing 40–80 kg. Simulation using the PK/PD model developed in this study may provide useful information for determining the analgesic effects and timing of the dosing interval after PTZ administration in perioperative patients.

Inhaler devices play an important role in the management of obstructive lung diseases including asthma, chronic obstructive pulmonary disease (COPD), and asthma–COPD overlap. Some of these patients show suboptimal inhaler techniques; however, time for inhaler instruction by pharmacists is limited in daily clinical practice. Therefore, sufficient education regarding inhaler device handling should be provided within a limited time frame. The current study aimed to investigate the instruction methods provided by community pharmacists and their influence on inhaler device handling techniques in outpatient clinical care settings. We retrospectively collected the data of outpatients with obstructive lung diseases who were referred to our hospital and who underwent inhalation technique assessments conducted by community pharmacists. The prevalence of handling errors, clinical characteristics of patients, and instruction methods were analyzed. In total, 138 patients (170 devices) were included in this study. Approximately 70.0% of patients received verbal explanations combined with leaflets about inhaler instructions. In a device-based analysis, 63 (37.1%) of 170 devices had at least one technical error and 18 (10.6%) of the devices had critical errors. Patients without critical errors received practical demonstration instructions from pharmacists combined with leaflets and verbal explanations more frequently than those with critical errors (22.8 vs. 0%, p < 0.01). This study revealed that patients with obstructive lung diseases commonly present with inhaler device handling errors and critical errors were observed with non-negligible frequency in daily practice in Japan. Combined instruction with leaflet, verbal explanation, and pharmacist demonstration may be effective in improving proper inhaler treatment.

Solid tumors habitually harbor regions with insufficient oxygen away from vasculature. Hypoxia is an important factor that confers malignant phenotypes like chemoresistance to tumor cells. We have demonstrated that cathepsin G (CG) stimulates cell aggregation in breast cancer MCF-7 cells by activating insulin-like growth factor-1 signaling. We investigated whether cancer cell aggregates induced by CG acquire hypoxia-dependent chemoresistance. Pimonidazole staining and hypoxia-inducible factor (HIF)-1α and -2α expression indicated that the core of the cell aggregates was hypoxic. Electrophoretic mobility shift and reporter assays showed that the CG-induced cell aggregates displayed transcriptional activity through HIF-responsive elements. Moreover, HIF target genes PGK1 and SLC2A1 demonstrated upregulated expression in CG-induced cell aggregates, indicating that the aggregates expressed functional HIF. Doxorubicin (DXR)-induced cytotoxicity was significantly lower in the cell aggregates induced by CG compared with monolayer cells under normoxia. Unexpectedly, the upregulation of P-glycoprotein expression, which is reported to be a HIF-target gene, and decreasing intracellular accumulation of DXR was not detected in the cell aggregates as opposed to in monolayer cells under normoxia. Additionally, reduction of DXR sensitivity in the aggregates was not suppressed by treatment with the HIF inhibitor, YC-1 and HIF-1α small interfering RNA (siRNA). Therefore, we conclude that cell aggregation induced by CG decreases DXR sensitivity via a HIF-independent mechanism.

trans-Banglene and cis(c)-banglene possess neurotrophin-like activity in rat neurons. However, the molecular mechanisms underlying t-banglene-induced neurotrophic activity in rat and human neurons remain unclear. Here, we performed transcriptome analysis in PC12 cells, a rat adrenal gland pheochromocytoma cell line treated with t-banglene, using comprehensive RNA sequencing. The differentially expressed gene analysis of the sequencing data revealed that the expression of RT1 class I, locus CE1 (RT1-CE1) was upregulated, and that of ATP binding cassette subfamily A member 1 (abca1), myosin light chain 6, and hippocampus abundant transcript 1 was downregulated in t-banglene-treated PC12 cells, with statistically significant differences. We also confirmed the RT1-CE1 upregulation and abca1 downregulation in t-banglene-treated PC12 cells by real-time reverse transcription quantitative PCR. RT1-CEl is a major histocompatibility complex class I (MHCI) protein. ABCAl is a major cholesterol transporter that regulates efflux of intracellular cholesterol and phospholipids. Thus, our results suggest an exciting link between MHCI, cholesterol regulation, and neural development.

Neudesin is a secretory protein involved in the brain development during embryonic period and diet-induced development of adipose tissue. Although neudesin is also expressed in the testis, its physiological functions in the testis have not been documented. Therefore, we examined neudesin-encoding neuron-derived neurotrophic factor (Nenf) gene-knockout (Neudesin-KO) mice to clarify the functions of neudesin in the testis. The testicular size of the Neudesin-KO mice was significantly smaller than that of wild-type (WT) mice. However, histological analyses did not reveal any abnormalities in the testis, caput epididymis, and cauda epididymis. Sperm number in the cauda epididymis was comparable between WT and KO mice. Neudesin-KO male mice produced vaginal plugs on paired WT female mice, with a frequency similar to that in WT male mice. A similar number of embryos were developed in the females copulated with WT and Neudesin-KO males. Molecular analysis indicated that the ion transporters Slc19a1 and Kcnk3 were more expressed in the testis of Neudesin-KO mice than in the testis of WT mice, suggesting that the transport of ions and some nutrients in the testis has some abnormalities. Testicular size decreased on postnatal day 6, but not on the day of birth, indicating that neudesin is involved in the postnatal, but not embryonic, development of testis. These results indicate a novel role of neudesin in the development of testis.

OX40, a member of the tumor necrosis factor (TNF) receptor superfamily, is induced on activated T cells. Membrane-bound OX40 ligand (OX40L) expressed by activated antigen-presenting cells induces OX40 signaling, which promotes T cell immunity. OX40 agonism would be a potential target for immunotherapy, however, it remains unclear how the activity of OX40 can be successfully controlled by a designer OX40L protein. We prepared a soluble OX40L protein possessing a PA-peptide tag and a collagenous trimerization domain from mannose-binding lectin (MBL), and tested whether PA-MBL-OX40L fusion protein worked as an agonist for OX40. We found that the majority of recombinant PA-MBL-OX40L protein purified from culture supernatants displayed a trimer structure and bound to cell surface OX40 or OX40-Fc fusion protein in a dose-dependent manner. Upon stimulation of CD4⁺ T cells with TCR/CD3 without CD28, PA-MBL-OX40L displayed significantly increased proliferative and cytokine responses when compared with a benchmark agonistic monoclonal antibody for OX40. Both soluble and immobilized forms of PA-MBL-OX40L induced potent OX40 signaling in CD4⁺ T cells. Mice administered with PA-MBL-OX40L displayed significantly augmented T cell-mediated delayed-type hypersensitivity responses. Our results suggest that activity of OX40L could be engineered to elicit better T cell responses by rational design of its assembly and architecture.

Bevacizumab is an inhibitor of vascular endothelial growth factor (VEGF) that prevents tumor growth. While bevacizumab is therapeutically effective, it induces several adverse events. Among these, central nervous system (CNS) ischemia can lead to death or permanent disability. In this study, we reviewed the Japanese Adverse Drug Event Report database to analyze the occurrence of CNS ischemia after bevacizumab administration. Significant associations between the occurrence of CNS ischemia and bevacizumab use were detected (adjusted reporting odds ratios (ROR): 2.68, 95% confidence interval (CI): 2.00–3.59, p < 0.001). Furthermore, an association between diagnosis of glioma and bevacizumab use was also detected (p < 0.001). These events occurred early after the start of treatment and then gradually decreased; however, more than half of CNS ischemia events were reported beyond 30 d after the first administration. In addition, a logistic regression suggested that CNS ischemia caused by bevacizumab was associated with glioma, underlying hypertension and aging. A poor prognosis was reported for several cases occurring in elderly patients (over 60 years of age). Although bevacizumab is a useful pharmacological treatment for cancer, caution should be taken to avoid severe adverse events. Accordingly, the patient’s general and medical condition should be carefully examined before initiating treatment, and blood pressure should be continuously assessed throughout treatment with bevacizumab to prevent CNS ischemia.

Human metabolic profiles for substances such as toxic food-derived compounds are usually allometrically extrapolated from traditionally determined in vivo rat concentration profiles. To evaluate internal exposures in humans without any reference to experimental data, physiologically based pharmacokinetic (PBPK) modeling could be used if the model input parameters could be estimated in silico. This approach would simplify the use of PBPK models for forward dosimetry after oral doses. In this study, the in silico estimation of input parameters for PBPK models (i.e., fraction absorbed × intestinal availability, absorption rate constants, and volumes of the systemic circulation) was updated for an panel of 355 chemicals (212 previously analyzed and 143 additional substances) using a light gradient boosting machine learning algorithms (LightGBM) based on between 11 and 29 in silico-calculated chemical descriptors. Simplified human PBPK models were then used to calculate virtual maximum plasma concentrations (C_max) and areas under the concentration–time curve (AUC) based on two sets of input parameters, i.e., traditionally derived values from in vivo data and those calculated in silico using the current updated systems. Both sets of C_max and AUC data were well correlated (r = 0.87 and r = 0.73, respectively; p < 0.01, n = 355). Therefore, input parameters for human PBPK models for a diverse range of compounds could be successfully estimated using chemical descriptors and in silico tools. This approach to pharmacokinetic modeling has potential for application in computational toxicology and in the clinical setting for assessing the potential risk of general chemicals.

Tamoxifen, which is used to treat advanced gynecological tumors, has been associated with tumor cell metastasis. Herein, we investigated the effect of tamoxifen on epithelial–mesenchymal transition in endometrial cancer and the associated signaling mechanism. Wound healing and invasion chamber assays, respectively, were performed to determine the migrative capacity and invasiveness of tamoxifen-stimulated endometrial carcinoma (RL95-2) cells. Western blotting and immunofluorescence were used to evaluate the expression of vimentin, E-cadherin, calpain 10 (CANP10), and neuropilin-1 (NRP1). Transfection of a CAPN10-harboring plasmid was used to overexpress CANP10 in RL95-2 cells, and small interfering RNAs were used to silence CANP10 and NRP1 expression. Tamoxifen induced migration, invasion, and morphological changes in RL95-2 cells. It also downregulated E-cadherin expression and upregulated vimentin, CANP10, and NRP1 expression. CANP10 silencing inhibited tamoxifen-induced NRP1 upregulation, and CANP10 or NRP1 silencing inhibited the migration and invasion of RL95-2 cells. CANP10 overexpression upregulated vimentin expression and downregulated that of E-cadherin and also increased cell migration and invasion. Silencing NRP1 protein expression inhibited the induction effect of CANP10 overexpression. In conclusion, tamoxifen promotes the epithelial–mesenchymal transition of RL95-2 cells via the CANP10/NRP1 signaling pathway. Thus, targeting CANP10 or NRP1 may be a novel strategy for preventing tamoxifen-induced endometrial cancer metastasis.

Endothelial dysfunction contributes to cardiometabolic disorders, including hypertension, obesity, and type 2 diabetes. Esaxerenone is a selective, nonsteroidal, high-affinity mineralocorticoid receptor blocker recently approved in Japan for the treatment of hypertension. Although imbalanced signaling between vasorelaxant and vasocontractile factors induced by endothelial stimulation is often observed in type 2 diabetic vessels, the effects of esaxerenone on endothelium-dependent responses in type 2 diabetes remain unclear. The aim of this study was to investigate the effect of esaxerenone on endothelium-dependent responses in superior mesenteric arteries isolated from type 2 diabetic Goto-Kakizaki (GK) rats. It was found that esaxerenone (3 mg/kg/d for 4 weeks, per os (p.o.)) partially ameliorated acetylcholine (ACh)-induced endothelium-derived hyperpolarizing factor (EDHF)-type relaxation and NS309, a potent activator of small- and intermediate-conductance Ca²⁺-activated K⁺ channels, -induced relaxation, and reduced ACh-induced endothelium-derived contracting factor (EDCF)-mediated contraction. These results suggest that esaxerenone ameliorates endothelial function through increased EDHF signaling and suppressed EDCF signaling.

SMTP-7, a fungal metabolite, is reported to have a high degree of availability for the ischemia–reperfusion (IR)-induced acute kidney injury (AKI) model. Cisplatin, a widely used anticancer drug, has serious side effects, such as AKI. Hence, we aimed to examine the effect of SMTP-7 on cisplatin-induced AKI in this study. Significant increases in blood urea nitrogen (BUN) and serum creatinine (Scr) were observed at 72 h after the intravenous infusion of cisplatin (20 mg/kg). Histologically, necrosis and dilatation (hyaline casts) as well as regeneration were observed in proximal tubules. SMTP-7 inhibited the elevation on BUN and Scr caused by cisplatin dose dependently. The efficacy of SMTP-7 was notable when the drug was administered on the day after cisplatin treatment, whereas the repeated administration of the drug did not result in an enhanced efficacy. Moreover, 10 mg/kg of SMTP-7 considerably ameliorated tubular necrosis and dilatation. The cisplatin treatment also caused an up-regulation of tumor necrosis factor-α (TNF-α) mRNA expression prior to the elevation of the levels of BUN and Scr. Administration of SMTP-7 (10 mg/kg) at 24 h after the cisplatin infusion alleviated the up-regulation of TNF-α mRNA expression. These findings suggest that SMTP-7 exhibits a renoprotective effect against cisplatin infusion based on the inhibition of the expression of pro-inflammatory cytokines such as TNF-α and may be expected a new effective drug for the treatment of cisplatin-induced AKI.

Phthalic acid (PA) diesters are widely used in consumer products, as plasticizers, and are ubiquitous environmental pollutants. There is a growing concern about their adjuvant effect on allergic diseases. Although its precise mechanism remains unknown, possible involvement of transient receptor potential ankyrin 1 (TRPA1) has been suggested. Hence, in this study, the activation of human and mouse TRPA1s by a series of PA di- and monoesters was investigated using a heterologous expression system in vitro. Consequently, it was found that monoesters activated human TRPA1, where EC₅₀ values were in the order of mono-hexyl > mono-heptyl > mono-n-octyl > mono-2-ethylhexyl > mono-isononyl and mono-isodecyl esters. Significant species differences in TRPA1 activation by PA monoesters were also discovered; PA monoesters activated human TRPA1 but not mouse TRPA1 in a concentration-dependent manner up to 50 µM. These findings suggest that PA esters may exert TRPA1-dependent adverse effects on humans, which have never been demonstrated in experimental animals.

Macrophages have important roles in the progression of inflammation. Ajania purpurea Shih. is a member of the Ajania Poljakor family that grows in Tibet (China). Extracts from plants in this genus have anti-bacterial and anti-inflammatory properties. However, there are few reports on the activity and mechanism of Ajania purpurea. Here, we confirmed the anti-inflammatory effect of Ajania purpurea Shih. ethanol extract (EAPS) by examining the levels of inflammatory factors in a mouse model of peritonitis and RAW264.7 cells. The main components of EAPS detected by LC-MS analysis included piperine and chlorogenic acid. In particular, in lipopolysaccharide (LPS)-induced RAW264.7 cells, EAPS inhibited the protein expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) in LPS-induced RAW264.7 cells, lowered the levels of nitric oxide (NO) and prostaglandin E2 (PGE₂), as well as the release of inflammatory factors such as tumor necrosis factor-alpha (TNF-α) and pro-inflammatory cytokines such as interleukin (IL)-1β and IL-6. In addition, Western blot analysis and immunofluorescence staining verified that EAPS inhibited the activity of the nuclear factor-kappaB (NF-κB) pathway by reducing the nuclear translocation of the p65 subunit. Furthermore, in a mouse model of peritonitis, EAPS inhibited the release of inflammatory factors, as well as the recruitment of immune cells including neutrophils and macrophages. These findings indicated that EAPS suppressed LPS-induced inflammation via inhibiting the NF-κB pathway in RAW264.7 cells and mice with peritonitis. Thus, EAPS may be a viable therapeutic method for the treatment of inflammation and related disorders.

Coronavirus disease 2019 (COVID-19), caused by the severe acute respiratory syndrome coronavirus 2 (SARS-COV-2) virus, has spread worldwide since 2019. Several studies report that adult patients hospitalized with severe COVID-19 can be treated successfully with remdesivir (RDV). However, few studies report the safety and efficacy of RDV for children. This study included 20 hospitalized patients who were diagnosed with COVID-19 and received RDV at Kobe Children’s Hospital, Japan, between February and June, 2022. The median age was 2 years (IQR, 1–11 years; range, 5 months to 19 years). Twelve (60%) patients were male. Three (15%) patients were previously healthy, whereas the other 17 had at least one underlying medical condition: five (25%) patients had respiratory disease, four (20%) had cardiac disease, three (15%) had central nervous system disease, four (20%) had hematologic or oncologic disease, and two (10%) had a chromosomal abnormal. All patients recovered without any sequelae, and no serious adverse events were reported. The adverse events were elevated liver enzymes in 4 children (20%), leukopenia (5%), neutropenia (5%), and hypokalemia (5%). Our study may show that the use of RDV for COVID-19 in children led to no serious adverse events.

Various side effects associated with the use of nonsteroidal anti-inflammatory drugs (NSAIDs) in analgesia have been reported. Among the NSAIDs, celecoxib has fewer side effects and is often used in therapeutic applications. However, the effect of celecoxib on aged skin is unknown. In this study, we investigated the effects of celecoxib administration on the skin of aged mice. We analyzed a 40-week-old mouse model and a 10-week-old mouse as the control group. The animals were orally administered celecoxib for four consecutive days and then killed and dissected the day after the last dose. In aged mice treated with celecoxib, the water content of the stratum corneum, which is one of the markers of dry skin, was lower than that in the control and young mice groups. In addition, serum hyaluronic acid, creatinine, and inflammatory cytokines in the collected blood samples of aged mice were elevated compared to those in other mice groups, suggesting the onset of acute renal injury. Therefore, it was considered that acute renal injury occurred from the administration of celecoxib to aged mice, whereas dry skin developed by the promotion of inflammatory cytokine secretion and release into the bloodstream in this group.