Mpk1 MAPK association with the Paf1 complex blocks Sen1-mediated premature transcription termination.

The Mpk1 MAPK of the yeast cell wall integrity pathway uses a noncatalytic mechanism to activate transcription of stress-induced genes by recruitment of initiation factors to target promoters. We show here that Mpk1 additionally serves a function in transcription elongation that is also independent of its catalytic activity. This function is mediated by an interaction between Mpk1 and the Paf1 subunit of the Paf1C elongation complex. A mutation in Paf1 that blocks this interaction causes a specific defect in transcription elongation of an Mpk1-induced gene, which results from Sen1-dependent premature termination through a Nab3-binding site within the promoter-proximal region of the gene. Our findings reveal a regulatory mechanism in which Mpk1 overcomes transcriptional attenuation by blocking recruitment of the Sen1-Nrd1-Nab3 termination complex to the elongating polymerase. Finally, we demonstrate that this mechanism is conserved in an interaction between the human ERK5 MAPK and human Paf1.

A Bee Trp-Arg Dense Peptide with Antiproliferation Efficacy against the Prostate Cancer Cell Line DU145.

Prostate cancer accounts for 14% of male cancer-related fatalities in the UK. Given the challenges associated with hormone-based therapies in the context of androgen-independent prostate cancer, there is an imperative need for research into anticancer drugs. N0821, a peptide belonging to the Trp-Arg dense region and derived from the homologous region of various bee species, shows substantial potential for an anticancer effect. Both MTT assays and 3D spheroid assays were conducted to substantiate its antiproliferation potential and strongly indicated the antiproliferation effect of N0820 (WWWWRWWRKI) and N0821 (YWWWWRWWRKI). Notably, the mechanism underlying this effect is related to the downregulation of CCNA2 and the upregulation of CCNE1. Cell cycle arrest results from the reduction of CCNA2 in the S/G2 phase, leading to the accumulation of CCNE1. Our peptides were predicted to make an α-helix structure. This can act as an ion channel in the cell membrane. Therefore, we analyzed genes implicated in the influx of calcium ions into the mitochondria. Trp-Arg dense-region peptides are known for their antibacterial properties in targeting cell membranes, making the development of resistance less likely. Hence, further research in this area is essential and promising.

Identification of a Novel ERK5 (MAPK7) Inhibitor, MHJ-627, and Verification of Its Potent Anticancer Efficacy in Cervical Cancer HeLa Cells.

Extracellular signal-regulated kinase 5 (ERK5), a member of the mitogen-activated protein kinase (MAPK) family, is involved in key cellular processes. However, overexpression and upregulation of ERK5 have been reported in various cancers, and ERK5 is associated with almost every biological characteristic of cancer cells. Accordingly, ERK5 has become a novel target for the development of anticancer drugs as inhibition of ERK5 shows suppressive effects of the deleterious properties of cancer cells. Herein, we report the synthesis and identification of a novel ERK5 inhibitor, MHJ-627, and verify its potent anticancer efficacy in a yeast model and the cervical cancer HeLa cell line. MHJ-627 successfully inhibited the kinase activity of ERK5 (IC50: 0.91 µM) and promoted the mRNA expression of tumor suppressors and anti-metastatic genes. Moreover, we observed significant cancer cell death, accompanied by a reduction in mRNA levels of the cell proliferation marker, proliferating cell nuclear antigen (PCNA), following ERK5 inhibition due to MHJ-627 treatment. We expect this finding to serve as a lead compound for further identification of inhibitors for ERK5-directed novel approaches for oncotherapy with increased specificity.

Filipendula glaberrima Nakai extract inhibits the bacterial infection by induction of HBD2 and HBD3 expression, and reduction of the inflammatory activity.

Defensins and inflammation are innate immune barriers of the body against infectious pathogens. Searching for a compound that can inhibit infectious diseases by affecting human ß-defensin (HBD) and proinflammatory cytokines is the new trend in research to control bacterial infection. The aim of this study is to provide a natural compound, Filipendula glaberrima Nakai extract (FGE), which is able to induce the expression of an antimicrobial defensin as well as reduce inflammation. FGE induced the expression of HBD2 and HBD3 through activating both p38 and NF-κB signaling pathways. Furthermore, FGE inhibited the expression of TNF-α and IL-6 via p38 and NF-κB pathways in Staphylococcus aureus-stimulated THP1 cells. Injection of FGE alleviated cutaneous erythema and swelling caused by S. aureus injection in mice ears. Taken together, FGE could reduce bacterial infection by inducing the expression of defensin and anti-inflammatory activity.

Alpinia japonica extract induces apoptosis of hepatocellular carcinoma cells through G0/G1 cell cycle arrest and activation of JNK.

Hepatic cancer was the third most prevalent cause of cancer-related death worldwide in 2018, and its incidence is increasing. While therapeutic agents for hepatic cancer have improved, these agents can cause serious side effects, including damage to healthy tissues. To overcome this limitation, more than 3,000 plants have been used globally as common alternatives for cancer treatment. The anti-cancer activity of Alpinia japonica, one of the traditional herbal medicines (Korean name: Kkot-yang-ha), was investigated. Water extract of A. japonica (AJ) decreased the cell viability of hepatic cancer cells. AJ extract showed greater than 70% loss of mitochondrial potential in HepG2 cells as demonstrated by JC-1 staining. Apoptosis was induced by treatment with AJ extract as shown through FACS analysis, and G0/G1 phase arrest of 76.66% HepG2 cells was confirmed through cell cycle analysis and quantitative RT-PCR. Improper regulation of ERK1/2 might contribute to cell death, and JNK activation is necessary for apoptosis induced by stress stimuli. AJ extract stimulated the phosphorylation of JNK and ERK1/2, mitogen-activated protein kinases (MAPKs), in HepG2 cells. AJ extract has anticancer activity by inhibiting cell cycle progression, leading to apoptosis of hepatic cancer cells. This extract could potentially be used as a therapeutic agent for hepatic cancer.

MSF Enhances Human Antimicrobial Peptide ß-Defensin (HBD2 and HBD3) Expression and Attenuates Inflammation via the NF-κB and p38 Signaling Pathways.

Both defensin and inflammation are part of the human innate immune system that responds rapidly to pathogens. The combination of defensins with pro- or anti-inflammatory effects can be a potential research direction for the treatment of infection by pathogens. This study aimed to identify whether MSF (Miracle Synergy material made using Filipendula glaberrima), a probiotic lysate of Filipendula glaberrima extracts fermented with Lactiplantibacillus plantarum K8, activates the expression of human ß-defensin (HBD2 and HBD3) to protect the host against pathogens and inhibit inflammation caused by S. aureus, in vitro with Western blot analysis, qRT-PCR and in vivo studies with a mouse model were used to evaluate the effects of MSF. The MSF treatment induced HBD2 and HBD3 expression via the p38 and NF-κB pathways. Furthermore, MSF treatment significantly reduced the expression of pro-inflammatory cytokines (TNF-α, IL-1ß, IL-6, and IL-8), also through p38 and NF-κB in S. aureus-induced inflammatory condition. MSF treatment remarkably reduced erythema in mice ears caused by the injection of S. aureus, while K8 lysate treatment did not initiate a strong recovery. Taken together, MSF induced the expression of HBD2 and HDB3 and activated anti-inflammatory activity more than the probiotic lysates of L. plantarum K8. These findings show that MSF is a potential defensin inducer and anti-inflammatory agent.

Development of in vitro three-dimensional drug screening system for obesity-related metabolic syndrome.

Metabolic syndrome is increasingly common, and closely related with overweight or obesity. In the obese state, macrophages infiltrate to the adipose tissue (AT), resulting in chronic inflammation and insulin resistance in the AT cells. Recently, attention has been paid to the role of AT macrophages in metabolic disorders should be applied to the initial drug screening step, but it was difficult to mimic the inflammatory adipocytes using the traditional 2-dimensional (2D) culture. In this study, we developed the 3-dimensional (3D) culture system to overcome this limitation. After adipogenic differentiation, lipid droplets were highly accumulated in cells, and differentiation of preadipocytes was not declined by macrophage co-culture. However, only co-cultured cells expressed the insulin resistance features. Compare to mono-cultured adipocytes, co-cultured adipocytes showed reduced glucose uptake and GLUT4 did not translocated to cell membrane even though treatment of high concentration of insulin. Using 3D co-culture model, we develop a microwell-scale drug screening protocol to test anti-obesity effect. 3D cultured cells reacted more sensitive to drugs, and PPARγ antagonist GW9662 (10, 20 µM) repressed adipogenic differentiation in a concentration-dependent manner in 3D co-cultured cells.

Evaluation of Dose-Dependent Obesity and Diabetes-Related Complications of Water Chestnut (Fruit of Trapa japonica) Extracts in Type II Obese Diabetic Mice Induced by 45% Kcal High-Fat Diet.

Background and Objectives: The currently used pharmacological agents for metabolic disorders such as type II diabetes have several limitations and adverse effects; thus, there is a need for alternative therapeutic drugs and health functional foods. Materials and Methods: This study investigated the pharmacological effects of water chestnut (fruit of Trapa japonica) extracts (WC: 50-200 mg/kg) for type II diabetes using a 45% Kcal high-fat diet (HFD)-fed type II obese diabetic mice model for a period of 84 days, and the effects were compared to those of metformin (250 mg/kg). Results: Increases in body weight, serum biochemical indices such as triglycerides, low-density lipoprotein, and blood urea nitrogen, increases in antioxidant defense system enzymes such as catalase, superoxide dismutase, and glutathione, and mRNA expressions (such as AMPKα1 and AMPKα2) in the liver tissue and mRNA expressions (such as AMPKα2 mRNA, leptin, and C/EBPα) in the adipose tissue were observed in the HFD control group. The WC (50 mg/kg)-administered group showed no significant improvements in diabetic complications. However, HFD-induced obesity and diabetes-related complications such as hyperlipidemia, diabetic nephropathy, nonalcoholic fatty liver disease (NAFLD), oxidative stress, activity of antioxidant defense systems, and gene expressions were significantly and dose-dependently inhibited and/or normalized by oral administration of WC (100 mg/kg and 200 mg/kg), particularly at a dose of 100 mg/kg. Conclusions: The results of this study suggest that WC at an appropriate dose could be used to develop an effective therapeutic drug or functional food for type II diabetes and various associated complications, including NAFLD.

Synthesis of novel 1H-Pyrazolo[3,4-b]pyridine derivatives as DYRK 1A/1B inhibitors.

As DYRK1A and 1B inhibitors, 1H-pyrazolo[3,4-b]pyridine derivatives were synthesized. Mostly, 3-aryl-5-arylamino compounds (6) and 3,5-diaryl compounds (8 and 9) were prepared and especially, 3,5-diaryl compound 8 and 9 showed excellent DYRK1B inhibitory enzymatic activities with IC50 Values of 3-287 nM. Among them, 3-(4-hydroxyphenyl), 5-(3,4-dihydroxyphenyl)-1H-pyrazolo[3,4-b]pyridine (8h) exhibited the highest inhibitory enzymatic activity (IC50 = 3 nM) and cell proliferation inhibitory activity (IC50 = 1.6 µM) towards HCT116 colon cancer cells. Also compound 8h has excellent inhibitory activities in patient-derived colon cancer organoids model as well as in 3D spheroid assay model of SW480 and SW620. The docking study supported that we confirmed that compound 8h binds to DYRK1B through various hydrogen bonding interactions and hydrophobic interactions.

Adenophora triphylla var. japonica Inhibits Candida Biofilm Formation, Increases Susceptibility to Antifungal Agents and Reduces Infection.

(1) Background: Candida is the most common cause of fungal infections worldwide, but due to the limited option of antifungal therapies, alternative strategies are required. (2) Methods: Adenophora triphylla var. japonica extract was used for the biofilm formation assay using RPMI1640. The combinatorial antifungal assay, the dimorphic transition assay, and the adherence assay were done to see the influence of inhibition of biofilm formation. qRT-PCR analysis were performed to check the gene expression. (3) Results: Adenophora triphylla var. japonica extract inhibited the Candida biofilm formation. Treatment of extract increased the antifungal susceptibility of miconazole from a 37% reduction in fungal growth to 99.05%, and also dose-dependently reduced the dimorphic transition of Candida and the attachment of Candida to HaCaT cells. The extract blocked the expression of hyphal-related genes, extracellular matrix genes, Ras1-cAMP-PKA pathway genes, Cph2-Tec1 pathway gene, and MAP kinase pathway gene. (4) Conclusions: In this study, the treatment of Adenophora triphylla var. japonica extract showed inhibition of fungal biofilm formation, activation of antifungal susceptibility, and reduction of infection. These results suggest that fungal biofilm formation is a good target for the development of antifungal adjuvants, and Adenophora triphylla var. japonica extract should be a good candidate for biofilm-associated fungal infections.

Development of a three-dimensional in vitro co-culture model to increase drug selectivity for humans.

AIM: Insulin resistance is a metabolic state where insulin sensitivity is lower than normal condition and strongly related to type 2 diabetes. However, an in vitro model mimicking insulin resistance is rare and thus screening drugs for insulin resistance severely depends on an in vivo model. Here, to increase anti-diabetic drug selectivity for humans, 3D ADMSCs and macrophages were co-cultured with in-house fabricated co-culture plates. MATERIAL AND METHODS: 3D co-culture plates were designed to load ADMSCs and RAW264.7 cells containing hydrogels in separate wells while allowing cell-cell interaction with co-culturing media. Hydrogels were constructed using a 3D cell-printing system containing 20 mg/ml alginate, 0.5 mg/ml gelatin and 0.5 mg/ml type I collagen. Cells containing hydrogels in 3D co-culture plates were incubated for 10 min to allow stabilization before the experiment. 3D co-culture plates were incubated with the CaCl2 solution for 5 min to complete the cross linking of alginate hydrogel. Cells in 3D co-culture plates were cultured for up to 12 days depending on the experiment and wells containing adipocytes and macrophages were separated and used for assays. RESULTS: KR-1, KR-2 and KR-3 compounds were applied during differentiation (12 days) in 3D co-cultured mouse 3T3-L1 adipocytes and 3D co-cultured human ADMSCs. Glucose uptake assay using 2-DG6P and 2-NBDG and western blot analysis were performed to investigate changes of insulin resistance in the 3D co-cultured model for interspecies selectivity of drug screening. KR-1 (mouse potent enantiomer) and KR-3 (racemic mixture) showed improvement of 2-DG and 2-NBDG uptake compared with KR-2 (human potent enantiomer) in 3D co-cultured 3T3-L1 adipocytes. In connection with insulin resistance in a 3D 3T3-L1 co-cultured model, KR-1 and KR-3 showed improvement of insulin sensitivity compared to KR-2 by markedly increasing GLUT4 expression. In contrast to the result of 3D co-cultured 3T3-L1 adipocytes, KR-1 failed to significantly improve 2-DG and 2-NBDG uptake in 3D co-cultured ADMSC adipocytes. Results of 2-NBDG accumulation and western blot analysis also showed that KR-2 and KR-3 improved insulin sensitivity relatively better than KR-1. CONCLUSIONS: Our 3D co-culture model with/without 3D co-culture plates can successfully mimic insulin resistance while allowing investigation of the effects of anti-obesity or anti-diabetic drugs on human or mouse co-culturing cell type. This 3D co-culture system may accelerate screening of drugs for insulin resistance depending on species.

Optimization of cyclic sulfamide derivatives as 11ß-hydroxysteroid dehydrogenase 1 inhibitors for the potential treatment of ischemic brain injury.

The 11ß-hydroxysteroiddehydrogenase type 1(11ß-HSD1), acortisolregenerating enzyme that amplifies tissue glucocorticoidlevels, plays an important role in diabetes, obesity, and glaucoma and is recognized as a potential therapeutic target for various disease conditions. Moreover, a recent study demonstrated that selective 11ß-HSD1 inhibitor can attenuate ischemic brain injury. This prompted us to optimize cyclic sulfamide derivative for aiming to treat ischemic brain injury. Among the synthesized compounds, 6e has an excellent in vitro activivity with an IC50 value of 1 nM toward human and mouse 11ß-HSD1 and showed good 11ß-HSD1 inhibition in ex vivo study using brain tissue isolated from mice. Furthermore, in the transient middle cerebral artery occlusion model in mice, 6e treatment significantly attenuated infarct volume and neurological deficit following cerebral ischemia/reperfusion injury. Additionally, binding modes of 6e for human and mouse 11ß-HSD1 were suggested.

Gentisic Acid Stimulates Keratinocyte Proliferation through ERK1/2 Phosphorylation.

Keratinocyte proliferation is important for skin wound healing. The wound healing process includes blood clotting around the wound, removal of dead cells and pathogens through inflammation, and then re-epithelialization through proliferation and maturation. Proliferation assay was performed on acid natural compounds to identify candidates for natural-derived components of skin injury treatment. We found that gentisic acid promoted high cell proliferation activity compared with other compounds. Gentisic acid improved HaCaT cell proliferation by over 20% in MTT assay. Gentisic acid also had higher healing activity in an in vitro wound healing assay than allantoin as a positive control. Furthermore, we have identified how the treatment of gentisic acid can increase proliferation in the cell. Western blot analysis of proteins in the mitogen-activated protein (MAP) kinase signaling pathway showed that ERK1/2 phosphorylation was increased by gentisic acid treatment. Thus, our study indicates that gentisic acid promotes the proliferation of keratinocyte by phosphorylation of ERK1/2.

A Novel Selective 11ß-HSD1 Inhibitor, (E)-4-(2-(6-(2,6-Dichloro-4-(Trifluoromethyl)Phenyl)-4-Methyl-1,1-Dioxido-1,2,6-Thiadiazinan-2-yl)Acetamido)Adamantan-1-Carboxamide (KR-67607), Prevents BAC-Induced Dry Eye Syndrome.

Dry eye syndrome is the most common eye disease and it is caused by various reasons. As the balance of the tear film that protects the eyes is broken due to various causes, it becomes impossible to properly protect the eyes. In this study, the protective effects and underlying mechanisms of topical (E)-4-(2-(6-(2,6-dichloro-4-(trifluoromethyl)phenyl)-4-methyl-1,1-dioxido-1,2,6-thiadiazinan-2-yl)acetamido)adamantan-1-carboxamide (KR-67607), a novel selective 11ß-hydroxysteroid dehydrogenase 1 (11ß-HSD1) inhibitor, were investigated in benzalkonium chloride (BAC)-induced dry eye syndrome. BAC-treated rat eyes induced significant increases in ocular surface damage, decreased corneal thickness, corneal basement membrane destruction in the conjunctival epithelium, and expression of pro-inflammatory cytokines tumor necrosis factor-α and 11ß-HSD1. These effects of BAC were reversed by topical KR-67607 treatment. Furthermore, KR-67607 decreased 4-hydroxynonenal expression and increased antioxidant and mucus secretion in BAC-treated rat eyes. Taken together, a novel selective 11ß-HSD1 inhibitor can prevent BAC-induced dry eye syndrome by inhibiting pro-inflammatory cytokine and reactive oxygen species expression via the inhibition of both 11ß-HSD1 activity and expression.

Toxicity Assessment of SiO2 and TiO2 in Normal Colon Cells, In Vivo and in Human Colon Organoids.

We conducted systemic assessments on the toxicity of silicon dioxide (SiO2) and titanium dioxide (TiO2) nanoparticles using different forms of normal colon cells (CCD-18Co), in vivo and in human colon organoids. The in vivo acute oral toxicity data showed that the LD50 values are greater than 2000 mg/kg for both the SiO2 and TiO2 nanoparticles; however, the SiO2 and TiO2 nanoparticles induced cytotoxicity in two-dimensional CCD-18Co cells and three-dimensional CCD-18Co spheroids and human colon organoids, with IC50 values of 0.6, 0.8 and 0.3 mM for SiO2 and 2.5, 1.1 and 12.5 mM for TiO2 nanoparticles, respectively. The data suggest that, when SiO2 and TiO2 are in nanoparticle form, cytotoxicity is induced; thus, care should be taken with these materials.

Cyp7a1 is continuously increased with disrupted Fxr-mediated feedback inhibition in hypercholesterolemic TALLYHO/Jng mice.

TALLYHO/Jng (TH) mice reveal hypercholesterolemia at an early age before their plasma glucose levels have increased. The increased plasma cholesterol should be related to bile acids (BAs) metabolism, because cholesterol is the precursor of BAs and BAs change cholesterol metabolism in a feedback manner. We analyzed the BAs pool size, BAs composition, and expression levels of several proteins that have key roles in BAs synthesis, excretion, and reabsorption and compared them to those of C57BL/6 (B6) mice to study BAs metabolism in TH mice. TH mice exhibited an increased total BAs pool size, increased BAs content in the cecum feces, and an increased ratio of muricholic acid (MCA)/cholic acid (CA). The mRNA and protein levels of cholesterol 7 alpha-hydroxylase (Cyp7a1) and the ATP-binding cassette sub-family G member 5 (Abcg5) were elevated in the liver but not in the apical sodium bile acid transporter (Asbt) and organic solute transporters (Osts) in the ileum. These results indicate that synthesis and the excretion of BAs from the liver to the gallbladder might be elevated, but the reabsorption rate of BAs in the ileum might be reduced. The declined expression of fibroblast growth factor 15 (Fgf15) and fibroblast growth factor receptor 4 (Fgfr4) was respectively identified in the ileum and the liver, indicating the disrupted feedback inhibition of Cyp7a1. Consequently, hypercholesterolemia in TH mice might increase the BAs amounts via the interrupted Fxr/Fgf15/Fgfr4-mediated feedback regulation of Cyp7a1.

Long-term functional voice outcomes after thyroidectomy, and effect of endotracheal intubation on voice.

PURPOSE: The aim of this study was to evaluate long-term functional voice outcomes after thyroidectomy, and the effect of endotracheal intubation on post-operative voice impairment. METHODS: We prospectively analyzed the voice outcomes of 155 serial thyroidectomy patients for up to 2 years. The control group consisted of 69 patients who underwent parotidectomy. Patients with post-operative recurrent laryngeal nerve palsy or palsy of the external branch of the superior laryngeal nerve were excluded. Self-assessment voice symptom scores (VSS) by questionnaire, and objective acoustic parameters and maximum phonation times, were evaluated pre-operatively and 1 day, 3 days, 1 week, 1 month, 3, 6, 12, 18, and 24 months after surgery. RESULTS: VSS increased from day 1 after surgery in both groups, and returned to pre-operative levels by 24 months in the thyroidectomy group and after 1 week post-operatively in the parotidectomy (control) group. The post-operative VSS of the thyroidectomy group was significantly higher than that of the control group up to 12 months post-operatively. Highest frequency decreased immediately after surgery in the thyroidectomy group, and recovered to pre-operative levels by 12 and 18 months in females and males, respectively, whereas it recovered to pre-operative levels after the first week in the control group. CONCLUSION: Impairment of voice function may persist for more than 18 months after thyroidectomy even in patients without RLN palsy. Endotracheal intubation can affect voice outcomes adversely for 1 week post-operatively.

Antifungal activity of magnoflorine against Candida strains.

Candida albicans is a major invasive pathogen, and the development of strains resistant to conventional antifungal agents has been reported in recent years. We evaluated the antifungal activity of 44 compounds against Candida strains. Magnoflorine showed the highest growth inhibitory activity of the tested Candida strains, with a minimum inhibitory concentration (MIC) of 50 µg/mL based on microdilution antifungal susceptibility testing. Disk diffusion assay confirmed the antifungal activity of magnoflorine and revealed that this activity was stable over 3 days compared to those of berberine and cinnamaldehyde. Cytotoxicity testing showed that magnoflorine could potentially be used in a clinical setting because it didn't have any toxicity to HaCaT cells even in 200 µg/mL of treatment. Magnoflorine at 50 µg/mL inhibited 55.91 ± 7.17% of alpha-glucosidase activity which is required for normal cell wall composition and virulence of Candida albicans. Magnoflorine also reduced the formation of C. albicans' biofilm. Combined treatment with magnoflorine and miconazole decreased the amount of miconazole required to kill various Candida albicans. Therefore, magnoflorine is a good candidate lead compound for novel antifungal agents.

Protective effects of carbenoxolone, an 11ß-HSD1 inhibitor, against chemical induced dry eye syndrome.

Dry eye syndrome (DES) is a disorder of the eye due to tear deficiency or excessive evaporation that causes damage to the eye and is associated with discomfort and dryness. 11ß-Hydroxysteroid dehydrogenase 1 (11ß-HSD1) is an enzyme that converts inactive cortisone to active cortisol. Recently, 11ß-HSD1 has been expressed in human and rodent eyes and has been recognized as a target of glaucoma. In this study, the therapeutic effects and underlying mechanisms of topical carbenoxolone, an 11ß-HSD1 inhibitor, were investigated in benzalkonium chloride (BAC)-treated human conjunctival epithelial cells and a rat DES model. In the in vitro study, carbenoxolone dose-dependently inhibited cell death and 11ß-HSD1 activity in BAC-treated human conjunctival epithelial cells. For the in vivo study, carbenoxolone or a solvent was administered to the BAC-induced DES model twice daily. BAC-treated rat eyes showed significant increases in ocular surface damage, a reduction of tears, decrease corneal thickness, corneal basement membrane destruction, apoptosis in the conjunctival epithelium, and expression of pro-inflammatory cytokines (TNF-α and IL-6) and 11ß-HSD1. These effects of BAC were reversed by topical carbenoxolone treatment. These results demonstrate that carbenoxolone can prevent DES by inhibiting pro-inflammatory cytokine expression and cell death of the corneal and conjunctival epithelium via inhibition of both 11ß-HSD1 activity and expression in the eyes of BAC-treated rats. It is suggested that topical 11ß-HSD1 inhibitors may provide a new therapeutic window in the prevention and/or treatment of DES.

Methionine sulfoxide reductase A deficiency exacerbates acute liver injury induced by acetaminophen.

Acetaminophen (APAP) overdose induces acute liver injury via enhanced oxidative stress and glutathione (GSH) depletion. Methionine sulfoxide reductase A (MsrA) acts as a reactive oxygen species scavenger by catalyzing the cyclic reduction of methionine-S-sulfoxide. Herein, we investigated the protective role of MsrA against APAP-induced liver damage using MsrA gene-deleted mice (MsrA-/-). We found that MsrA-/- mice were more susceptible to APAP-induced acute liver injury than wild-type mice (MsrA+/+). The central lobule area of the MsrA-/- liver was more impaired with necrotic lesions. Serum alanine transaminase, aspartate transaminase, and lactate dehydrogenase levels were significantly higher in MsrA-/- than in MsrA+/+ mice after APAP challenge. Deletion of MsrA enhanced APAP-induced hepatic GSH depletion and oxidative stress, leading to increased susceptibility to APAP-induced liver injury in MsrA-deficient mice. APAP challenge increased Nrf2 activation more profoundly in MsrA-/- than in MsrA+/+ livers. Expression and nuclear accumulation of Nrf2 and its target gene expression were significantly elevated in MsrA-/- than in MsrA+/+ livers after APAP challenge. Taken together, our results demonstrate that MsrA protects the liver from APAP-induced toxicity. The data provided herein constitute the first in vivo evidence of the involvement of MsrA in hepatic function under APAP challenge.