Casein Kinase 2 Alpha Inhibition Protects against Sepsis-Induced Acute Kidney Injury.

Sepsis-induced acute kidney injury (AKI) is a common complication in critically ill patients, often resulting in high rates of morbidity and mortality. Previous studies have demonstrated the effectiveness of casein kinase 2 alpha (CK2α) inhibition in ameliorating ischemia-reperfusion-induced AKI. In this study, our aim was to investigate the potential of the selective CK2α inhibitor, 4,5,6,7-tetrabromobenzotriazole (TBBt), in the context of sepsis-induced AKI. To assess this, we initially confirmed an upregulation of CK2α expression following a cecum ligation and puncture (CLP) procedure in mice. Subsequently, TBBt was administered to a group of mice prior to CLP, and their outcomes were compared to those of sham mice. The results revealed that, following CLP, the mice exhibited typical sepsis-associated patterns of AKI, characterized by reduced renal function (evidenced by elevated blood urea nitrogen and creatinine levels), renal damage, and inflammation (indicated by increased tubular injury score, pro-inflammatory cytokine levels, and apoptosis index). However, mice treated with TBBt demonstrated fewer of these changes, and their renal function and architecture remained comparable to that of the sham mice. The anti-inflammatory and anti-apoptotic properties of TBBt are believed to be associated with the inactivation of the mitogen-activated protein kinase (MAPK) and nuclear factor κB (NF-κB) signaling pathways. In conclusion, these findings suggest that inhibiting CK2α could be a promising therapeutic strategy for treating sepsis-induced AKI.

Aqueous extract of Chrysanthemum morifolium Ramat. inhibits RANKL-induced osteoclast differentiation by suppressing the c-fos/NFATc1 pathway.

OBJECTIVE: The flower of chrysanthemum, used worldwide as a medicinal and edible product, has shown various bioactivities, such as anti-inflammatory, antioxidant, anti-tumorigenic, and hepatoprotective activities, as well as cardiovascular protection. However, the effect of Chrysanthemum morifolium Ramat. on the regulation of osteoclast differentiation has not yet been reported. In this study, we aimed to investigate the inhibitory effect of Chrysanthemum morifolium Ramat. water extract (CME) on RANKL-induced osteoclast differentiation in mouse bone marrow-derived macrophages (BMMs). STUDY DESIGN: Bone marrow-derived macrophages (BMMs) isolated from the C57BL/6 J mice. The viability of BMMs was detected with MTT assays. Inhibitory effects of CME on osteoclast differentiation and bone resorption was measured by TRAP staining and Pit assay. Osteoclast differentiation-associated gene expression were assessed by Real-time quantitative polymerase chain reaction. Intracellular signaling molecules was assessed by western blot. RESULTS: CME significantly inhibited osteoclast differentiation in BMMs without cytotoxicity, besides inhibiting MAPK/c-fos and PLCγ2/CREB activation. The inhibitory effects of CME on differentiation-related signaling molecules resulted in significant repression of NFATc1 expression, which is a key transcription factor in osteoclast differentiation, fusion, and activation. CONCLUSION: Our results confirmed the inhibition of RANKL-induced PLCγ2/CREB/c-fos/NFATc1 activation by CME during osteoclast differentiation. The findings collectively suggested CME as a traditional therapeutic agent for osteoporosis, RA, and periodontitis.

ß-Cyclodextrin/Triclosan Complex-Grafted Methacrylated Glycol Chitosan Hydorgel by Photocrosslinking via Visible Light Irradiation for a Tissue Bio-Adhesive.

Accelerating wound healing with minimized bacterial infection has become a topic of interest in the development of the new generation of tissue bio-adhesives. In this study, we fabricated a hydrogel system (MGC-g-CD-ic-TCS) consisting of triclosan (TCS)-complexed beta-cyclodextrin (ß-CD)-conjugated methacrylated glycol chitosan (MGC) as an antibacterial tissue adhesive. Proton nuclear magnetic resonance (1H NMR) and differential scanning calorimetry (DSC) results showed the inclusion complex formation between MGC-g-CD and TCS. The increase of storage modulus (G') of MGC-g-CD-ic-TCS after visible light irradiation for 200 s indicated its hydrogelation. The swollen hydrogel in aqueous solution resulted in two release behaviors of an initial burst and sustained release. Importantly, in vitro and in vivo results indicated that MGC-g-CD-ic-TCS inhibited bacterial infection and improved wound healing, suggesting its high potential application as an antibacterial tissue bio-adhesive.

Myeloid cell-specific sirtuin 6 deficiency delays wound healing in mice by modulating inflammation and macrophage phenotypes.

We recently reported that myeloid cell-expressed sirtuin 6 (Sirt6) plays a crucial role in M1 macrophage polarization and chemotaxis. Given the prominent role of macrophages during wound repair and macrophage heterogeneity, we hypothesized that a Sirt6 deficiency in myeloid cells would delay skin wound closure by affecting the phenotypes of macrophages in wounds. To address this question, a full-thickness excisional lesion was made in the dorsal skin of myeloid cell-specific Sirt6 knockout (KO) and wild-type mice. Wound closure was delayed in the KO mice, which exhibited less collagen deposition, suppressed angiogenesis, and reduced expression of wound healing-related genes compared to the wild-type mice. Using immunohistochemical, flow cytometric, and gene-expression analyses of macrophage subpopulations from wound tissue, we identified increased infiltration of M1 macrophages with a concomitant decrease in M2 macrophage numbers in the KO mice compared to the wild-type mice. Consistent with the in vivo wound closure defects observed in the KO mice, keratinocytes and fibroblasts treated with KO macrophage-derived conditioned medium migrated slower than those treated with wild-type macrophage-derived conditioned medium. An analysis of downstream signaling pathways indicated that impaired Akt signaling underlies the decreased M2 phenotypic switching in KO mice. These results suggest that a macrophage phenotypic switch induced by Sirt6 deficiency contributes to impaired wound healing in mice.

Atopic dermatitis-like skin lesions are suppressed in fat-1 transgenic mice through the inhibition of inflammasomes.

Previous clinical trials have addressed the beneficial effects of fish oil supplementation on atopic dermatitis. Recently, we reported that fat-1 mice, which can convert n-6 to n-3 polyunsaturated fatty acids (PUFAs), are protected against allergic airway inflammation because their Th2 immune responses are suppressed. Here, we examined the effects of endogenously synthesized n-3 PUFAs on atopic dermatitis, a representative Th2-dominant allergic inflammatory disease. Mouse models of atopic dermatitis-like skin lesions were prepared by epicutaneous application of 2,4-dinitrochlorobenzene (DNCB) or house dust mite (HDM) extract to the ears. DNCB-treated fat-1 mice exhibited markedly reduced epidermal thickening, lower mast cell infiltration, and lower serum IgE and histamine compared with wild-type mice. The draining lymph nodes of fat-1 mice were substantially smaller and contained significantly smaller proportions of activated CD4+ T cells and IL-4-producing Th2 cells than those of wild-type mice. Consistent with these findings, the mRNA levels of Th2 cytokines were significantly decreased in DNCB-sensitized skin lesions of fat-1 mice. Lastly, inflammasome activation, IL-1ß production, and pyroptotic cell injury were suppressed in fat-1 mice. Similar results were observed in HDM-challenged fat-1 mice. This study confirms the results of previous clinical studies and suggests fish oil supplementation as a therapeutic strategy for atopic dermatitis-like skin lesions.

SPA0355 attenuates ischemia/reperfusion-induced liver injury in mice.

Hepatic ischemia/reperfusion (I/R) injury leads to oxidative stress and acute inflammatory responses that cause liver damage and have a considerable impact on the postoperative outcome. Much research has been performed to develop possible protective techniques. We aimed to investigate the efficacy of SPA0355, a synthetic thiourea analog, in an animal model of hepatic I/R injury. Male C57BL/6 mice underwent normothermic partial liver ischemia for 45 min followed by varying periods of reperfusion. The animals were divided into three groups: sham operated, I/R and SPA0355 pretreated. Pretreatment with SPA0355 protected against hepatic I/R injury, as indicated by the decreased levels of serum aminotransferase and reduced parenchymal necrosis and apoptosis. Liver synthetic function was also restored by SPA0355 as reflected by the prolonged prothrombin time. To gain insight into the mechanism involved in this protection, we measured the activity of nuclear factor-κB (NF-κB), which revealed that SPA0355 suppressed the nuclear translocation and DNA binding of NF-κB subunits. Concomitantly, the expression of NF-κB target genes such as IL-1ß, IL-6, TNF-α and iNOS was significantly downregulated. Lastly, the liver antioxidant enzymes superoxide dismutase, catalase and glutathione were upregulated by SPA0355 treatment, which correlated with the reduction in serum malondialdehyde. Our results suggest that SPA0355 pretreatment prior to I/R injury could be an effective method to reduce liver damage.

Guggulsterone inhibits melanogenesis in B16 murine melanoma cells by downregulating tyrosinase expression.

In the present study, we investigated the effect of guggulsterone on melanogenesis in B16 melanoma cells and elucidated its possible mechanism of action. The effects of guggulsterone on melanogenesis were determined by assaying melanin synthesis and cellular tyrosinase activity in B16/F10 mouse melanoma cells. Guggulsterone dose-dependently inhibited isobutylmethylxanthine (IBMX)-induced melanogenesis and cellular tyrosinase activity with no cytotoxicity. Decreased melanin biosynthesis was accompanied by the reduced expression of melanogenesis-related genes, such as tyrosinase, microphthalmia-associated transcription factor, tyrosinase-related protein (TRP)-1 and TRP-2. Guggulsterone also inhibited α-melanocyte stimulating hormone- or forskolin-induced increases in melanogenesis, suggesting an action on the cAMP-dependent melanogenic pathway. Co-incubation with chenodeoxycholic acid, a well-known farnesoid-X receptor agonist, did not affect IBMX-induced melanogenesis. These results suggest that guggulsterone exerts a melanogenic inhibitory effect through the downregulation of tyrosinase expression.

Saponified evening primrose oil reduces melanogenesis in B16 melanoma cells and reduces UV-induced skin pigmentation in humans.

This study was conducted to determine whether saponified evening primrose oil (sap-EPO) has the potential for use as a whitening agent and to investigate its underlying mechanisms of action. In B16 melanoma cells, sap-EPO dose-dependently inhibited isobutylmethylxanthine-induced melanogenesis with no cytotoxicity. This decrease in melanin production was correlated with reduced enzyme activity and decreased mRNA and protein levels of tyrosinase. The mRNA levels of tyrosinase-related proteins 1 and 2 decreased in response to treatment with sap-EPO, indicating that it regulated tyrosinase at the transcriptional level. Expression of microphthalmia-associated transcription factor was also decreased by sap-EPO as evidenced by decreased mRNA and protein levels. Additionally, topical application of sap-EPO resulted in efficient whitening of UVB-induced hyperpigmentation of human skin. Taken together, these results suggest that sap-EPO has the potential for use as a cosmetic whitening agent.

Saponified sunflower and safflower oils inhibit melanogenesis in B16 melanoma cells.

The purpose of this study was to determine whether saponified sunflower and safflower oils can potentially be used as whitening agents, and to investigate their underlying mechanisms. Saponified sunflower (sap-SU) and safflower (sap-SA) oils dose-dependently inhibited isobutylmethylxanthine-induced melanogenesis in B16 melanoma cells, with no cytotoxicity. This decrease in melanin production was correlated with reduced enzyme activity and decreased mRNA and protein levels of tyrosinase. mRNA levels of microphthalmia-associated transcription factor and tyrosinase-related proteins 1 and 2 were also decreased by sap-SU and sap-SA, indicating the regulation of tyrosinase at the transcriptional level. Taken together, these results suggest that sap-SU and sap-SA can potentially be used as cosmetic whitening agents.

Effect of xanthohumol on melanogenesis in B16 melanoma cells.

Xanthohumol (XH), the principal prenylflavonoid of the hop plant (Humulus lupulus L.), dose-dependently inhibited isobutylmethylxanthine (IBMX)-induced melanogenesis in B16 melanoma cells, with little cytotoxicity at the effective concentrations. Decreased melanin content was accompanied by reduced tyrosinase enzyme activity, protein and mRNA expression. The levels of tyrosinase-related protein 1 and 2 mRNAs were decreased by XH. XH also inhibited alpha-melanocyte stimulating hormone- or forskolin-induced increases in melanogenesis, suggesting an action on the cAMP-dependent melanogenic pathway. XH downregulated the protein and mRNA expression of microphthalmia-associated transcription factor (MITF), a master transcriptional regulator of key melanogenic enzymes. These results suggest that XH might act as a hypo-pigmenting agent through the downregulation of MITF in the cAMP-dependent melanogenic pathway.

Effect of Angelica gigas extract on melanogenesis in B16 melanoma cells.

During the screening of herbs for inhibition of melanogenesis, it was observed that ethanolic extract of Angelicae Gigantis Radix (AGE) effectively inhibited isobutylmethylxanthine-induced melanogenesis in B16 melanoma cells. The melanin content was significantly decreased by AGE in a dose-dependent manner, and no cytotoxicity was observed at the effective concentrations. Decreased melanin content was accompanied by reduced enzyme activity as well as reduced expression of tyrosinase protein and mRNA. The level of tyrosinase-related protein 1 and 2 mRNAs was also decreased by AGE. Additionally, AGE effectively inhibited alpha-melanocyte stimulating hormone- and forskolin-induced melanogenesis, and downregulated the mRNA expression of microphthalmia-associated transcription factor, a master transcriptional regulator of melanogenic genes. These results suggest that AGE acts as a putative hypopigmenting agent through downregulation of tyrosinase expression induced via a cAMP-dependent pathway.

Effect of scopoletin on lipoprotein lipase activity in 3T3-L1 adipocytes.

Hypertriglyceridemia is an independent risk factor of cardiovascular diseases. It is caused by the imbalance between hepatic triglyceride production and peripheral removal. Lipoprotein lipase (LPL) plays a central role in the removal of plasma triglyceride. During the screening of possible anti-dyslipidemic drugs, we observed that scopoletin (6-methoxy-7-hydroxycoumarin) significantly increased LPL activity in adipocytes. Scopoletin increased LPL activity in culture medium of 3T3-L1 adipocytes in dose- and time-dependent manners. It did not release LPL from the adipocyte membrane and, instead, increased the LPL mRNA level, suggesting transcriptional control. Scopoletin also partially reversed tumor necrosis factor-alpha-induced suppression of LPL activity. These results suggest the possible action of scopoletin as a facilitator of plasma triglyceride clearance.

Diallyl disulfide accelerates adipogenesis in 3T3-L1 cells.

Adipocyte differentiation is regulated by the sequential activation of transcription factors such as the CAAT/enhancer binding protein and peroxisome proliferator- activated receptor-gamma (PPAR-gamma). Several recent studies have shown that regulators of chromatin structure are also involved in adipocyte differentiation. Here we investigated the effects of diallyl disulfide (DADS), an oil-soluble sulfur compound found in processed garlic and an inhibitor of histone deacetylase (HDAC), on adipogenesis. Treatment with DADS accelerated terminal differentiation of 3T3-L1 cells into adipocytes as evidenced by Oil red O staining and cellular triglyceride assay results. Notably, the inhibition of HDAC during the first 2 days was sufficient to stimulate adipogenesis. Western blot analysis revealed that DADS increased the level of acetylated histones H3 and H4. In addition, DADS increased the expression of adipogenesis-related genes; LPL, FAS, SREBP1c, aP2 and PPAR-gamma, and decreased the expression of pref-1, a preadipocyte marker gene. Taken together, our results suggest that DADS affects adipocyte differentiation through histone acetylation at an early phase of adipocyte differentiation.

Stimulation of melanogenesis by scoparone in B16 melanoma cells.

AIM: The effect of coumarin derivatives on melanogenesis was investigated in B16 murine melanoma cells. METHODS: Melanin content and tyrosinase activity were analyzed spectrophotometrically. The expression of tyrosinase, tyrosinase-related protein-1 (TRP-1) and tyrosinase-related protein-2 (TRP-2) were measured either by reverse transcription-polymerase chain reaction (RT-PCR) or Western blot. RESULTS: Among the coumarin derivatives studied, scoparone (6,7- dimethoxycoumarin) was the most potent; the 6- or 7-methoxy group was found to be essential for the stimulation of melanogenesis. The melanin content was greatly increased by scoparone in a dose-dependent manner; there was no cytotoxicity at the effective concentrations. Scoparone increased enzyme activity as well as protein and mRNA expression of tyrosinase. In addition, mRNA of TRP-1 and TRP-2 were also increased after treatment with scoparone. H-89, an inhibitor of protein kinase A (PKA), completely inhibited the scoparone-induced increase of melanogenesis and the tyrosinase protein. CONCLUSION: These results suggest that scoparone-induced stimulation of melanogenesis is likely to occur at the transcriptional level of melanogenesis-related enzymes through PKA signaling.