Establishment of Intestinal Organoid from Rousettus leschenaultii and the Susceptibility to Bat-Associated Viruses, SARS-CoV-2 and Pteropine Orthoreovirus.

Various pathogens, such as Ebola virus, Marburg virus, Nipah virus, Hendra virus, Severe Acute Respiratory Syndrome Coronavirus (SARS-CoV), Middle East Respiratory Syndrome Coronavirus (MERS-CoV), and SARS-CoV-2, are threatening human health worldwide. The natural hosts of these pathogens are thought to be bats. The rousette bat, a megabat, is thought to be a natural reservoir of filoviruses, including Ebola and Marburg viruses. Additionally, the rousette bat showed a transient infection in the experimental inoculation of SARS-CoV-2. In the current study, we established and characterized intestinal organoids from Leschenault's rousette, Rousettus leschenaultii. The established organoids successfully recapitulated the characteristics of intestinal epithelial structure and morphology, and the appropriate supplements necessary for long-term stable culture were identified. The organoid showed susceptibility to Pteropine orthoreovirus (PRV) but not to SARS-CoV-2 in experimental inoculation. This is the first report of the establishment of an expandable organoid culture system of the rousette bat intestinal organoid and its sensitivity to bat-associated viruses, PRV and SARS-CoV-2. This organoid is a useful tool for the elucidation of tolerance mechanisms of the emerging rousette bat-associated viruses such as Ebola and Marburg virus.

Correction: Elbadawy, M., et al. Novel Functions of Death-Associated Protein Kinases through Mitogen-Activated Protein Kinase-Related Signals. Int. J. Mol. Sci. 2018, 19, 3031.

The authors wish to make the following corrections to this paper [...].

Establishment of a novel experimental model for muscle-invasive bladder cancer using a dog bladder cancer organoid culture.

In human and dogs, bladder cancer (BC) is the most common neoplasm affecting the urinary tract. Dog BC resembles human muscle-invasive BC in histopathological characteristics and gene expression profiles, and could be an important research model for this disease. Cancer patient-derived organoid culture can recapitulate organ structures and maintains the gene expression profiles of original tumor tissues. In a previous study, we generated dog prostate cancer organoids using urine samples, however dog BC organoids had never been produced. Therefore we aimed to generate dog BC organoids using urine samples and check their histopathological characteristics, drug sensitivity, and gene expression profiles. Organoids from individual BC dogs were successfully generated, expressed urothelial cell markers (CK7, CK20, and UPK3A) and exhibited tumorigenesis in vivo. In a cell viability assay, the response to combined treatment with a range of anticancer drugs (cisplatin, vinblastine, gemcitabine or piroxicam) was markedly different in each BC organoid. In RNA-sequencing analysis, expression levels of basal cell markers (CK5 and DSG3) and several novel genes (MMP28, CTSE, CNN3, TFPI2, COL17A1, and AGPAT4) were upregulated in BC organoids compared with normal bladder tissues or two-dimensional (2D) BC cell lines. These established dog BC organoids might be a useful tool, not only to determine suitable chemotherapy for BC diseased dogs but also to identify novel biomarkers in human muscle-invasive BC. In the present study, for the 1st time, dog BC organoids were generated and several specifically upregulated organoid genes were identified. Our data suggest that dog BC organoids might become a new tool to provide fresh insights into both dog BC therapy and diagnostic biomarkers.

Ovarian follicular changes and hemodynamics in Egyptian buffaloes under CIDR-PGF2α and Ovsynch-CIDR estrus synchronization treatments.

This study aimed at investigating the efficacy of two protocols of estrous synchronization on follicular changes and hemodynamics. Pluriparous Egyptian buffaloes (n = 36) were synchronized either with controlled internal drug release (CIDR)-PGF2α (7-days CIDR insert with PGF2α injected on the 6th day; n = 18) or Ovsynch-CIDR (Ovsynch protocol concurrent with 7-days CIDR insert; n = 18). Blood sampling and ovarian ultrasound examinations (Grayscale, color and power Doppler modes) were conducted on the Day of CIDR removal, estrus, and luteal phase. Mean follicle diameter (MFD), first (1st-LF) and second (2nd-LF) largest follicle diameters, and E2 levels significantly increased in the CIDR-PGF2α group at CIDR withdrawal. Ovsynch-CIDR markedly fortified higher follicle population, MFD, and 1st-LF diameter at estrus and corpus luteum (CL) volume at the luteal phase in concomitant with increases (P < 0.05) in E2 (at estrus) and P4 (at luteal phase). At CIDR removal, the blue pixels in the dominant follicle (DF) were higher (1.5 times; P = 0.054) in the Ovsynch-CIDR than in the CIDR-PGF2α. At estrus, total blood flow (TBF) and power Doppler pixels (PDP) to DF(s) were noticeably higher (seven and 1.6 times; respectively) in the Ovsynch-CIDR than in CIDR-PGF2α (5906 ± 237 vs. 830 ± 60 pixels, P < 0.01 and 5479 ± 322 vs. 3377 ± 19 pixels, P < 0.05; respectively). At the luteal phase, TBF and PDP to the CL increased in the Ovsynch-CIDR group than in the CIDR-PGF2α group (11060 ± 965 vs. 7963 ± 480 pixels, 1.4 times, P = 0.05 and 18900 ± 1350 vs. 13220 ± 568 pixels, 1.1 times, P = 0.005; respectively). In conclusion, based on the improvement in synchronized follicular activity and hemodynamics, the Ovsynch-CIDR regimen is persuaded in Egyptian buffaloes.

Emerging Roles of C-Myc in Cancer Stem Cell-Related Signaling and Resistance to Cancer Chemotherapy: A Potential Therapeutic Target Against Colorectal Cancer.

Myc is a nuclear transcription factor that mainly regulates cell growth, cell cycle, metabolism, and survival. Myc family proteins contain c-Myc, n-Myc, and l-Myc. Among them, c-Myc can become a promising therapeutic target molecule in cancer. Cancer stem cells (CSCs) are known to be responsible for the therapeutic resistance. In the previous study, we demonstrated that c-Myc mediates drug resistance of colorectal CSCs using a patient-derived primary three-dimensional (3D) organoid culture. In this review, we mainly focus on the roles of c-Myc-related signaling in the regulation of CSCs, chemotherapy resistance, and colorectal cancer organoids. Finally, we introduce the various types of c-Myc inhibitors and propose the possibility of c-Myc as a therapeutic target against colorectal cancer.

Influence of glucocorticoids on time-of-day-dependent variations in IgE-, histamine-, and platelet-activating factor-mediated systemic anaphylaxis in different mouse strains.

A time-of-day-dependent variation in IgE-mediated passive systemic anaphylaxis was previously reported in ICR mice. In the present study, we investigated time-of-day-dependent variations in IgE-, histamine-, and platelet-activating factor (PAF)-mediated systemic anaphylaxis in C57BL/6, BALB/c, and NC/Nga mice at 9:00 h and 21:00 h, and evaluated the potential influence of glucocorticoids (GCs) on these variations. We found significant time-of-day-dependent variations in IgE-mediated systemic anaphylaxis in C57BL/6 mice, and in histamine- and PAF-mediated systemic anaphylaxis in BALB/c mice. Significant daily variations in IgE-, histamine-, and PAF-mediated systemic anaphylaxis were not observed in NC/Nga mice. Pretreatment with dexamethasone and adrenalectomy abolished the daily variations in IgE-mediated systemic anaphylaxis in C57BL/6 mice and in PAF-mediated systemic anaphylaxis in BALB/c mice, suggesting that GCs from adrenal glands are pivotal in regulating these variations. In contrast, pretreatment with dexamethasone and adrenalectomy did not abolish the daily variation in histamine-mediated systemic anaphylaxis in BALB/c mice, suggesting that GC-independent and adrenal gland-independent mechanisms are important for the variation. The present study demonstrated that time-of-day-dependent variations in systemic anaphylaxis differed among inbred mouse strains and with anaphylaxis-inducing substances. Thus, mouse strains, time of experiment, and anaphylaxis-inducing substances used must be considered to obtain appropriate experimental results.

Novel Functions of Death-Associated Protein Kinases through Mitogen-Activated Protein Kinase-Related Signals.

Death associated protein kinase (DAPK) is a calcium/calmodulin-regulated serine/threonine kinase; its main function is to regulate cell death. DAPK family proteins consist of DAPK1, DAPK2, DAPK3, DAPK-related apoptosis-inducing protein kinases (DRAK)-1 and DRAK-2. In this review, we discuss the roles and regulatory mechanisms of DAPK family members and their relevance to diseases. Furthermore, a special focus is given to several reports describing cross-talks between DAPKs and mitogen-activated protein kinases (MAPK) family members in various pathologies. We also discuss small molecule inhibitors of DAPKs and their potential as therapeutic targets against human diseases.

Hedgehog Signals Mediate Anti-Cancer Drug Resistance in Three-Dimensional Primary Colorectal Cancer Organoid Culture.

Colorectal cancer is one of the most common causes of cancer death worldwide. In patients with metastatic colorectal cancer, combination treatment with several anti-cancer drugs is employed and improves overall survival in some patients. Nevertheless, most patients with metastatic disease are not cured owing to the drug resistance. Cancer stem cells are known to regulate resistance to chemotherapy. In the previous study, we established a novel three-dimensional organoid culture model from tumor colorectal tissues of human patients using an air-liquid interface (ALI) method, which contained numerous cancer stem cells and showed resistance to 5-fluorouracil (5-FU) and Irinotecan. Here, we investigate which inhibitor for stem cell-related signal improves the sensitivity for anti-cancer drug treatment in tumor ALI organoids. Treatment with Hedgehog signal inhibitors (AY9944, GANT61) decreases the cell viability of organoids compared with Notch (YO-01027, DAPT) and Wnt (WAV939, Wnt-C59) signal inhibitors. Combination treatment of AY9944 or GANT61 with 5-FU, Irinotecan or Oxaliplatin decreases the cell viability of tumor organoids compared with each anti-cancer drug alone treatment. Treatment with AY9944 or GANT61 inhibits expression of stem cell markers c-Myc, CD44 and Nanog, likely through the decrease of their transcription factor, GLI-1 expression. Combination treatment of AY9944 or GANT61 with 5-FU or Irinotecan also prevents colony formation of colorectal cancer cell lines HCT116 and SW480. These findings suggest that Hedgehog signals mediate anti-cancer drug resistance in colorectal tumor patient-derived ALI organoids and that the inhibitors are useful as a combinational therapeutic strategy against colorectal cancer.

Preplanned safety analysis of the JFMC37-0801 trial: a randomized phase III study of six months versus twelve months of capecitabine as adjuvant chemotherapy for stage III colon cancer.

BACKGROUND: Six months of adjuvant chemotherapy is regarded as the standard of care for patients with stage III colon cancer. However, whether longer treatment can improve prognosis has not been fully investigated. We conducted a phase III study comparing 6 and 12 months of adjuvant capecitabine chemotherapy for stage III colon cancer, and report here the results of our preplanned safety analysis. METHODS: Patients aged 20-79 years with curatively resected stage III colon cancer were randomly assigned to receive 8 cycles (6 months) or 16 cycles (12 months) of capecitabine (2500 mg/m2/day on days 1-14 of each 21-day cycle). Treatment exposure and adverse events (AEs) were evaluated. RESULTS: A total of 1304 patients (642 and 636 in the 6-month and 12-month groups, respectively) were analyzed. The most common AE was hand-foot syndrome (HFS). HFS, leukocytopenia, neutropenia, and hyperbilirubinemia (any grade) occurred more frequently in the 12-month group than in the 6-month group. HFS was the only grade ≥3 AE to have a significantly higher incidence in the 12-month group (23 vs 17%, p = 0.011). The completion rate for 8 cycles was 72% in both groups, while that for 16 cycles was 46% in the 12-month group. HFS was the most common AE requiring dose reduction and treatment discontinuation. CONCLUSIONS: Twelve months of adjuvant capecitabine demonstrated a higher cumulative incidence of HFS compared to the standard 6-month treatment period, while toxicities after 12 months of capecitabine were clinically acceptable. TRIAL REGISTRATION: UMIN-CTR, UMIN000001367.

Open-label, randomized, comparative, phase III study on effects of reducing steroid use in combination with Palonosetron.

The purpose of this study is to compare the efficacy of a single administration of dexamethasone (DEX) on day 1 against DEX administration on days 1-3 in combination with palonosetron (PALO), a second-generation 5-HT3 receptor antagonist, for chemotherapy-induced nausea and vomiting (CINV) in non-anthracycline and cyclophosphamide (AC) moderately-emetogenic chemotherapy (MEC). This phase III trial was conducted with a multi-center, randomized, open-label, non-inferiority design. Patients who received non-AC MEC as an initial chemotherapy were randomly assigned to either a group administered PALO (0.75 mg, i.v.) and DEX (9.9 mg, i.v.) prior to chemotherapy (study treatment group), or a group administered additional DEX (8 mg, i.v. or p.o.) on days 2-3 (control group). The primary endpoint was complete response (CR) rate. The CR rate difference was estimated by logistic regression with allocation factors as covariates. The non-inferiority margin was set at -15% (study treatment group - control group). From April 2011 to March 2013, 305 patients who received non-AC MEC were randomly allocated to one of two study groups. Overall, the CR rate was 66.2% in the study treatment group (N = 151) and 63.6% in the control group (N = 154). PALO plus DEX day 1 was non-inferior to PALO plus DEX days 1-3 (difference, 2.5%; 95% confidence interval [CI]: -7.8%-12.8%; P-value for non-inferiority test = 0.0004). There were no differences between the two groups in terms of complete control rate (64.9 vs 61.7%) and total control rate (49.7% vs 47.4%). Anti-emetic DEX administration on days 2-3 may be eliminated when used in combination with PALO in patients receiving non-AC MEC.

[Bacteria isolated from surgical infections and its susceptibilities to antimicrobial agents--special references to bacteria isolated between April 2010 and March 2011].

Bacteria isolated from surgical infections during the period from April 2010 to March 2011 were investigated in a multicenter study in Japan, and the following results were obtained. In this series, 631 strains including 25 strains of Candida spp. were isolated from 170 (81.7%) of 208 patients with surgical infections. Four hundred and twenty two strains were isolated from primary infections, and 184 strains were isolated from surgical site infection. From primary infections, anaerobic Gram-negative bacteria were predominant, followed by aerobic Gram-negative bacteria, while from surgical site infection aerobic Gram-positive bacteria were predominant, followed by anaerobic Gram-negative bacteria. Among aerobic Gram-positive bacteria, the isolation rate of Enterococcus spp. such as Enterococcus faecalis, Enterococcus faecium, and Enterococcus avium was highest, followed by Streptococcus spp. such as Streptococcus anginosus and Staphylococcus spp. such as Staphylococcus aureus, in this order, from primary infections, while Enterococcus spp. such as E. faecalis and E. faecium was highest, followed by Staphylococcus spp. such as S. aureus from surgical site infection. Among aerobic Gram-negative bacteria, Escherichia coli was the most predominantly isolated from primary infections, followed by Klebsiella pneumoniae, Klebsiella oxytoca, Enterobacter cloacae, and Pseudomonas aeruginosa in this order, and from surgical site infection, E. coli and R aeruginosa were most predominantly isolated, followed by E. cloacae and K. pneumoniae. Among anaerobic Gram-positive bacteria, the isolation rates of Parvimonas micra, Eggerthella lenta, Streptococcus constellatus, Gemella morbillorum, and Collinsella aerofaciens were the highest from primary infections, and the isolation rate from surgical site infection was generally low. Among anaerobic Gram-negative bacteria, the isolation rate of Bilophila wadsworthia was the highest from primary infections, followed by, Bacteroides fragilis and Bacteroides ovatus, and from surgical site infection, B. fragilis was most predominantly isolated, followed by Bacteroides thetaiotaomnicron, in this order. In this series, vancomycin-resistant MRSA (methicillin-resistant S. aureus), vancomycin-resistant Enterococcus spp. and multidrug-resistant P. aeruginosa were not observed.

[Bacteria isolated from surgical infections and its susceptibilities to antimicrobial agents - Special references to bacteria isolated between April 2011 and March 2012].

Bacteria isolated from surgical infections during the period from April 2011 to March 2012 were investigated in a multicenter study in Japan, and the following results were obtained. In this series, 785 strains including 31 strains of Candida spp. were isolated from 204 (78.8%) of 259 patients with surgical infections. Five hundred and twenty three strains were isolated from primary infections, and 231 strains were isolated from surgical site infection. From primary infections, anaerobic Gram-negative bacteria were predominant, followed by aerobic Gram-negative bacteria, while from surgical site infection aerobic Gram-positive bacteria were predominant, followed by anaerobic Gram-negative bacteria. Among aerobic Gram-positive bacteria, the isolation rate of Enterococcus spp. was highest, followed by Streptococcus spp. and Staphylococcus spp., in this order, from primary infections, while Enterococcus spp. was highest, followed by Staphylococcus spp. from surgical site infection. Among aerobic Gram-negative bacteria, Escherichia coli was the most predominantly isolated from primary infections, followed by Klebsiella pneumoniae, Pseudomonas aeruginosa and Enterobacter cloacae, in this order, and from surgical site infection, E. coli was most predominantly isolated, followed by P. aeruginosa, K. pneumoniae, and E. cloacae. Among anaerobic Gram-positive bacteria, the isolation rate of Eggerthella lenta was the highest from primary infections, followed by Parvimonas micra, Collinsella aerofaciens, Lactobacillus acidophilus and Finegoldia magna, and from surgical site infection, E. lenta was most predominantly isolated, followed by P micra and L. acidophilus, in this order. Among anaerobic Gram-negative bacteria, the isolation rate of Bacteroidesfragilis was the highest from primary infections, followed by Bilophila wadsworthia, Bacteroides thetaiotaomicron, Bacteroides uniformis and Bacteroides vulgatus, and from surgical site infection, B. fragilis was most predominantly isolated, followed by Bacteroides caccae, B. thetaiotaomicron, Bacteroides ovatus and B. wadsworthia, in this order. In this series, vancomycin-resistant MRSA (methicillin-resistant Staphylococcus aureus), vancomycin-resistant Enterococcus spp. and multidrug-resistant P. aeruginosa were not observed. We should carefully follow up B. wadsworthia which was resistant to various antimicrobial agents, and also Bacteroides spp. which was resistant to many ß-lactams.

Comparative pharmacokinetics of tyrosine kinase inhibitor, lapatinib, in dogs and cats following single oral administration.

Lapatinib is an orally administered tyrosine kinase inhibitor used to treat human epidermal growth factor receptor 2 (HER2) -overexpressing breast cancers in humans. Recently, the potential of lapatinib treatment against canine urothelial carcinoma or feline mammary tumor was investigated. However, the pharmacokinetic studies of lapatinib in dogs and cats are not well-defined. In the present study, the pharmacokinetic characteristics of lapatinib in both cats and dogs after a single oral administration at a dose of 25 mg/kg were compared with each other. Lapatinib was administered orally to four female laboratory cats and four female beagle dogs. Blood samples were collected over time, and the plasma lapatinib concentrations were analyzed by HPLC. Following a single dose of 25 mg/kg, the averaged maximum plasma concentration (Cmax) of lapatinib in cats was 0.47 µg/mL and achieved at 7.1 hr post-administration, while the Cmax in dogs was 1.63 µg/mL and achieved at 9.5 hr post-administration. The mean elimination half-life was 6.5 hr in cats and 7.8 hr in dogs. The average area under the plasma concentration-time curve of dogs (37.2 hr·µg/mL) was significantly higher than that of cats (7.97 hr·µg/mL). These results exhibited slow absorptions of lapatinib in both animals after oral administration. The Cmax observed in cats was significantly lower and the half-life was shorter than those observed in dogs. Based on these results, a larger dose or shorter dosing intervals might be recommended in cats to achieve similar plasma concentration as dogs.

Biliary excretion and pharmacokinetics of several fluoroquinolones after intravenous injection in rabbits.

The aim of this study was to measure the concentrations of enrofloxacin (ERFX) and other fluoroquinolones; orbifloxacin (OBFX), marbofloxacin (MBFX), and ofloxacin (OFLX) in the plasma and bile of rabbits after a single intravenous (IV) injection. Twenty male rabbits were divided into four groups and given each drug by IV injection into the ear vein at a dose of 5.0 mg/kg BW. The concentration of ERFX, ciprofloxacin (CPFX), OBFX, MBFX and OFLX in plasma and bile were determined by HPLC. CPFX, metabolite of ERFX, was also measured by HPLC in plasma and bile of rabbits receiving ERFX. Several pharmacokinetic parameters in plasma were calculated and biliary clearance (CLbile) was calculated from extent of biliary excretion and accumulation of AUC of each drug. After IV injection, elimination half-life (t1/2ß) was 4.13, 3.68, 6.60, 5.14 hr; volume of distribution at a steady state (Vdss) was 1.24, 0.503, 0.771, 1.02 L/kg; and total body clearance (CLtot) was 1.05, 0.418, 0.271, 0.453 L/kg/hr, respectively. The values for CLbile for ERFX, OBFX, MBFX, and OFLX were 0.0048, 0.0050, 0.0057, and 0.0094 L/kg/hr, respectively. These values represent 0.48%, 1.2%, 2.1%, and 2.3% of the total body clearance (CLtot) of each drug, respectively. The biliary clearance of CPFX was also measured and found to be 0.0199 L/kg/hr with ERFX administration. The results showed that ERFX, OBFX, MBFX, and OFLX were not excreted into the bile to a significant extent, making them safe drugs to use in rabbits.

Effect of xylazine sedation on testicular blood flow, testicular echotexture, and circulating hormones in Shiba goats.

Under field conditions, sedation may be required for a full assessment of the reproductive potential of farm animals. The present study aimed to investigate the effect of xylazine sedation on testicular hemodynamics (TBF), echotexture, testicular volume (TV), and circulating hormones in goats. Sixteen male Shiba goats were sedated using the recommended dose of xylazine (0.05 mg/Kg BW). Testicular hemodynamics were evaluated using color-pulsed Doppler ultrasonography before and after sedation. Echotexture of the testicular parenchyma and TV were assessed using computerized image analysis. Concentrations of testosterone, estradiol (E2), inhibin, cortisol, follicle-stimulating hormone (FSH), and luteinizing hormone (LH) were measured using radioimmunoassay. There were no effects of xylazine sedation in TBF, TV, testicular parenchyma parameters, and concentrations of testosterone, inhibin, FSH, and LH (P ˃ 0.05). However, after sedation, there was significantly (P ˂ 0.05) lower cortisol and E2 concentration (42.88 ± 6.79 ng/ml and 2.47 ± 0.58 pg/ml, respectively) than before sedation (94.89 ± 13.74 ng/ml and 8.65 ± 1.79 pg/ml, respectively). The required time to perform the full scanning of the testis was significantly lower (8.50 ± 0.38 min) after xylazine sedation compared to the non-sedated goats (25.75 ± 1.14 min). In conclusion, xylazine sedation may be practically recommended for the evaluation of TBF in goats because it did not significantly alter velocities parameters and Doppler indices of blood flow within the testicular arteries. Most plasma hormones did not significantly change; however, E2 and cortisol were significantly reduced after xylazine administration.

Anti-cancer activity of Chaga mushroom (Inonotus obliquus) against dog bladder cancer organoids.

Despite its disadvantages, chemotherapy is still commonly used for the treatment of bladder cancer (BC). Developing natural supplements that can target cancer stem cells (CSCs) which cause drug resistance and distant metastasis is necessary. Chaga mushrooms are popular to have several health-promoting and anti-cancer potentials. Organoid culture can recapitulate tumor heterogeneity, epithelial environment, and genetic and molecular imprints of the original tissues. In the previous study, we generated dog bladder cancer organoids (DBCO) as a novel experimental model of muscle-invasive BCO. Therefore, the present study aimed to examine the anti-tumor potentials of Chaga mushroom extract (Chaga) against DBCO. Four strains of DBCO were used in the present study. Treatment with Chaga inhibited the cell viability of DBCO in a concentration-dependent way. Treatment of DBCO with Chaga has significantly arrested its cell cycle and induced apoptosis. Expression of bladder CSC markers, CD44, C-MYC, SOX2, and YAP1, declined in the Chaga-treated DBCO. Also, Chaga inhibited the phosphorylation of ERK in DBCO. Expression of downstream signals of ERK, C-MYC, and Cyclins (Cyclin-A2, Cyclin-D1, Cyclin-E1, and CDK4) was also inhibited by Chaga in DBCO. Interestingly, the combinational treatment of DBCO with Chaga and anti-cancer drugs, vinblastine, mitoxantrone, or carboplatin, showed a potentiating activity. In vivo, Chaga administration decreased tumor growth and weight of DBCO-derived xenograft in mice with the induction of necrotic lesions. In conclusion, Chaga diminished the cell viability of DBCO by inhibiting proliferation-related signals and stemness conditions as well as by arresting the cell cycle. Collectively, these data suggest the value of Chaga as a promising natural supplement that could potentiate the effect of adjuvant chemotherapy, lower its adverse effects, and thus, limit the recurrence and metastasis of BC.

Oral pharmacokinetics of sulfadiazine and sulfamonomethoxine in female Holstein milking cows.

The efficacy of orally administered drugs in cattle is thought to be slow because of the anatomical and physiological features of their forestomach. Thus, parenteral routes are mainly preferred to administer drugs. However, the effect of some drugs with unique physicochemical properties was promptly obtained even after oral administration in clinically ill cattle. Therefore, the present study aimed to investigate pharmacokinetically the usefulness of the oral route in cattle by comparing the oral pharmacokinetic properties of two sulfonamides with different physicochemical properties. Sulfadiazine (SDZ) and sulfamonomethoxine (SMM) were administered by intravenous and oral route to four female Holstein cows with a 4-weeks washout period. Blood samples were collected over time, and SDZ and SMM concentrations in plasma were analyzed by HPLC. Data obtained from the same animal after intravenous and oral administration were simultaneously analyzed with the one compartment model, and kinetic parameters were calculated. The Tmax (mean ± SD) of SMM (2.75 ± 0.96 hr) was significantly achieved earlier than that of SDZ (5.00 ± 1.15 hr). Further, the mean absorption time of SMM (5.24 ± 0.69 hr) was significantly shorter than that of SDZ (5.92 ± 1.11 hr). Also, the half-life of absorption of SMM (3.91 ± 0.51 hr) was significantly shorter than that of SDZ (4.51 ± 0.82 hr). These data suggest that the absorption rates of highly unionized drugs (such as SMM) from the forestomach of cattle may be markedly higher than less unionized ones (such as SDZ).

Pharmacokinetics and 28-day repeated-dose toxicity of enniatin B after oral administration in mice.

Enniatins are emerging mycotoxins that contaminate foods. The present study investigated the oral pharmacokinetics and 28-day repeated-dose oral toxicity of enniatin B (ENNB) in CD1 (ICR) mice. In the pharmacokinetic study, male mice received a single oral or intravenous dose of ENNB [30 mg/kg body weight (BW) and 1 mg/kg BW, respectively]. After oral dosing, ENNB exhibited 139.9% bioavailability, a 5.1-h elimination half-life, 5.26% fecal excretion from 4 to 24 h post-dose, and upregulation of Cyp7a1, Cyp2a12, Cyp2b10, and Cyp26a1 in the liver 2 h post-dosing. In the 28-day toxicity study, ENNB was administered to male and female mice by oral gavage at 0, 7.5, 15, and 30 mg/kg BW/day. Females (7.5 and 30 mg/kg) showed dose-unrelated decreased food consumption without accompanying changes in clinical parameters. Males (30 mg/kg) showed low red blood cell counts and high blood urea nitrogen levels and absolute kidney weights; however, other related parameters including the histopathology of systemic organs/tissues were unchanged. These results suggest that ENNB may not induce toxicity after 28 days of oral administration in mice, despite high absorption. The no-observed-adverse-effect level of ENNB after 28 days of repeated oral doses was 30 mg/kg BW/day for both sexes of mice.

Evaluation of the efficacy of mitochondrial fission inhibitor (Mdivi-1) using non-alcoholic steatohepatitis (NASH) liver organoids.

Non-alcoholic steatohepatitis (NASH) is known to progress to cirrhosis and hepatocellular carcinoma in some patients. Although NASH is associated with abnormal mitochondrial function related to lipid metabolism, mechanisms for the development and effective treatments are still unclear. Therefore, new approaches to elucidate the pathophysiology are needed. In the previous study, we generated liver organoids from different stages of NASH model mice that could recapitulate the part of NASH pathology. In the present study, we investigated the relationship between mitochondrial function and NASH disease by comparing NASH liver organoids (NLO) and control liver organoids (CLO). Compared with CLO, mitochondrial and organoid morphology was abnormal in NLO, with increased expression of mitochondrial mitogen protein, DRP1, and mitochondria-derived reactive oxygen species (ROS) production. Treatment of NLO with a DPR1 inhibitor, Mdivi-1 resulted in the improvement of morphology and the decreased expression of fibrosis-related markers, Col1a1 and Acta2. In addition, treatment of NASH model mice with Mdivi-1 showed a decrease in fatty liver. Mdivi-1 treatment also prevented fibrosis and ROS production in the liver. These results indicate that NLO undergoes enhanced metabolism and abnormal mitochondrial morphology compared with CLO. It was also suggested that Mdivi-1 may be useful as a therapeutic agent to ameliorate NASH pathology.

Establishment of an experimental model of canine malignant mesothelioma organoid culture using a three-dimensional culture method.

Canine malignant mesothelioma (cMM) is a rare and drug-resistant malignant tumor. Due to few patients and experimental models, there have not been enough studies to demonstrate the pathogenesis of the disease and novel effective treatment for cMM. Since cMM resembles human MM (hMM) in histopathological characteristics, it is also considered a promising research model of hMM. Compared with conventional 2-dimensional (2D) culture methods, 3-dimensional (3D) organoid culture can recapitulate the properties of original tumor tissues. However, cMM organoids have never been developed. In the present study, we for the first time generated cMM organoids using the pleural effusion samples. Organoids from individual MM dogs were successfully generated. They exhibited the characteristics of MM and expressed mesothelial cell markers, such as WT-1 and mesothelin. The sensitivity to anti-cancer drugs was different in each strain of cMM organoids. RNA sequencing analysis showed cell adhesion molecule pathways were specifically upregulated in cMM organoids compared with their corresponding 2D cultured cells. Among these genes, the expression level of E-cadherin was drastically higher in the organoids than that in the 2D cells. In conclusion, our established cMM organoids might become a new experimental tool to provide new insights into canine and human MM therapy.