[Show abstract][Hide abstract] ABSTRACT: Cholangiopathy is a chronic immune-mediated disease of the liver, which is characterized by cholangitis, ductular reaction and biliary-type hepatic fibrosis. There is no proven medical therapy that changes the course of the disease. In previous studies, melittin was known for attenuation of hepatic injury, inflammation and hepatic fibrosis. This study investigated whether melittin provides inhibition on cholangitis and biliary fibrosis in vivo. Feeding 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) to mice is a well-established animal model to study cholangitis and biliary fibrosis. To investigate the effects of melittin on cholangiopathy, mice were fed with a 0.1% DDC-containing diet with or without melittin treatment for four weeks. Liver morphology, serum markers of liver injury, cholestasis markers for inflammation of liver, the degree of ductular reaction and the degree of liver fibrosis were compared between with or without melittin treatment DDC-fed mice. DDC feeding led to increased serum markers of hepatic injury, ductular reaction, induction of pro-inflammatory cytokines and biliary fibrosis. Interestingly, melittin treatment attenuated hepatic function markers, ductular reaction, the reactive phenotype of cholangiocytes and cholangitis and biliary fibrosis. Our data suggest that melittin treatment can be protective against chronic cholestatic disease in DDC-fed mice. Further studies on the anti-inflammatory capacity of melittin are warranted for targeted therapy in cholangiopathy.
[Show abstract][Hide abstract] ABSTRACT: Atherosclerosis is a complex disease which involves both genetic and environmental factors in its development and progression. Shear stress is the drag force per unit area acting on the endothelium as a result of blood flow, and it plays a critical role in plaque location and progression. TGF-β1 is often regarded to have pro-atherosclerotic effect on vascular disease. TGF-β1 downstream targets Smad, for regulating a set of genes associated with atherosclerosis. Therefore, modulation of TGF-β1 and Smad expression may be the important targets for the prevention and treatment of shear stress-induced vascular disease. However, the precise mechanism of the anti-atherosclerotic effects of novel therapeutic approach has not been elucidated by using animal models regarding the shear stress-induced vascular disease. Therefore, we designed to test whether Smad decoy ODN would prevent the development of atherosclerosis in the shear stress-induced ApoE-/-mice on a western diet. We examined the effect of cast placement on the development of atherosclerosis, and the carotid artery was harvested at the sacrifice to observe histological changes. Also, we evaluated the impact of Smad decoy ODN in the regulation of genes expression related to atherosclerosis, including TGF-β1, PAI-1, and α-SMA. Our results showed that western diet with cast placement developed atherosclerosis in ApoE-/-mouse. Also, administration of Smad decoy ODN decreases the expression of TGF-β1, PAI-1, and α-SMA. These results demonstrate the potential of Smad decoy ODN to prevent the progression of atherosclerosis in ApoE-/-mouse model with western diet and shear stress.
International journal of clinical and experimental pathology 06/2015; 8(4):3971-3978. · 1.89 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Propionibacterium acnes (P. acnes) cause inflammatory acne and play an important role in the pathogenesis of acne by inducing inflammatory mediators. P. acnes contributes to the inflammatory responses of acne by activating inflammatory cells, keratinocytes and sebocytes to secrete pro‑inflammatory cytokines such as tumor necrosis factor‑α (TNF‑α), interleukin (IL)‑1β and IL‑8. Bee venom has traditionally been used in the treatment of certain immune‑related diseases. However, there has not yet been a robust trial to prove the therapeutic effect of bee venom in skin inflammation. The aim of the present study was to investigate anti‑inflammatory properties of bee venom in skin inflammation induced by P. acnes using keratinocytes (HaCaT) and monocytes (THP‑1). P. acnes is known to stimulate the production of pro‑inflammatory cytokines such as IL‑1, IL‑8, IL‑12 and TNF‑α. In the present study, the production of interferon‑γ (IFN‑γ), IL‑1β, IL‑8 and TNF‑α was increased by P. acnes treatment in HaCaT and THP‑1 cells. By contrast, bee venom effectively inhibited the secretion of IFN‑γ, IL‑1β, IL‑8 and TNF‑α. Furthermore, P. acnes treatment activated the expression of IL‑8 and toll‑like receptor 2 (TLR2) in HaCaT cells. However, bee venom inhibited the expression of IL‑8 and TLR2 in heat‑killed P. acnes. Based on these results, it is concluded that bee venom has an effective anti‑inflammatory activity against P. acnes in HaCaT and THP‑1 cells. Therefore, we suggest that bee venom is an alternative treatment to antibiotic therapy of acne.
International Journal of Molecular Medicine 04/2015; 35(6). DOI:10.3892/ijmm.2015.2180 · 2.09 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Chronic liver diseases with different aetiologies rely on the chronic activation of liver injuries which result in a fibrogenesis progression to the end stage of cirrhosis and liver failure. Based on the underlying cellular and molecular mechanisms of a liver fibrosis, there has been proposed several kinds of approaches for the treatment of liver fibrosis. Recently, liver gene therapy has been developed as an alternative way to liver transplantation, which is the only effective therapy for chronic liver diseases. The activation of hepatic stellate cells, a subsequent release of inflammatory cytokines and an accumulation of extracellular matrix during the liver fibrogenesis are the major obstacles to the treatment of liver fibrosis. Several targeted strategies have been developed, such as antisense oligodeoxynucleotides, RNA interference and decoy oligodeoxynucleotides to overcome this barriers. With this report an overview will be provided of targeted strategies for the treatment of liver cirrhosis, and particularly, of the targeted gene therapy using short RNA and DNA segments.
World Journal of Gastroenterology 10/2014; 20(40):14696-14705. DOI:10.3748/wjg.v20.i40.14696 · 2.37 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Propionibacterium acnes (P. acnes) is a major contributing factor to the inflammatory component of acne. The many prescription medications for acne allow for a large number of potential combination treatments. However, several antibiotics, apart from their antibacterial effects, exert side‑effects, such as the suppression of host inflammatory responses. Purified bee venom (BV) is a natural toxin produced by honeybees (Apis mellifera L.). BV has been widely used as a traditional medicine for various diseases. In the present study, to investigate the therapeutic effects of BV against P. acnes-induced inflammatory skin disease, P. acnes was intradermally injected into the ears of mice. After the injection, BV was applied to the skin surface of the right ear. Histological observation revealed that P. acnes induced a considerable increase in the number of infiltrated inflammatory cells. However, treatment with BV markedly reduced these reactions compared with the P. acnes-injected mice not treated with BV. Moreover, the expression levels of tumor necrosis factor (TNF)-α, and interleukin (IL)-1β were significantly reduced in the BV-treated mice compared with the untreated P. acnes-injected mice. In addition, treatment with BV significantly inhibited Toll-like receptor (TLR)2 and CD14 expression in P. acnes-injected tissue. The binding activity of nuclear factor-κB (NF-κB) and activator protein (AP)-1 was markedly suppressed following treatment with BV. The results from our study, using an animal model, indicate that BV exerts an inhibitory effect on inflammatory skin diseases. In conclusion, our data indicate that BV has potential for use as an anti-acne agent and may be useful in the pharmaceutical and cosmetics industries.
International Journal of Molecular Medicine 09/2014; 34(5). DOI:10.3892/ijmm.2014.1933 · 2.09 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The appearance of proliferating bile ductular structures, which is called the "atypical ductular reaction" is frequently observed in various chronic liver diseases associated. However, the origin of these increased bile ductules has been a matter of controversy. In this study, we investigated the origin of ductular cells as an aspect of relation between epithelial to mesenchymal transition (EMT) and epithelial members of liver parenchyme, such as hepatocyte and cholangiocyte by immunohistochemical staining of human liver. Thirteen specimens of surgically resected liver with biliary cirrhosis were selected. Three sets of double immunohistochemical stains were done; Hep-Par 1 - cytokeratin 19 (CK19), Hep-Par 1 - α-sm ooth mus cle actin (α-SMA) and CK19 - α-SMA. As a result, we investigated the dual expression of the markers of hepatocyte and cholangiocyte in the same cell; in ductular cell and surrounding hepatocyte. However, there seems to be no dual expression of markers for EMT with epithelial markers. This study suggests a possibility of phenotypic change of mature hepatocyte into cholangiocyte. Future studies will be necessary to determine the role that proliferating cholangiocytes play in the pathogenesis of biliary fibrosis and how cholangiocytes interact with other cell types of the liver such as hepatic stellate cells or Kupffer cells.
International journal of clinical and experimental pathology 08/2014; 7(7):4076-85. · 1.89 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The tumor microenvironment has many roles involving tumor progression, invasion and metastasis. The tumor cells at the tumor border loose epithelial properties and acquire mesenchymal features. This, epithelial-to-mesenchymal transition (EMT) has been suggested to be an important process for tissue and lymphovascular invasion. Pulmonary tissue samples from 15 patients with primary adenocarcinoma were evaluated with using immunofluorescence multi-staining the EMT-associated markers including E-cadherin and alpha-smooth muscle actin (α-SMA), and transcription factors including E-SNAIL and SLUG, and ZEB1. The data were analyzed in specific area, such as tumor center and tumor border. In this study we show that the invasive adenocarcinoma differentially expressed SNAIL and SLUG, and Zeb1 and it was associated with the loss of epithelial marker (E-cadherin) and gaining of mesenchymal marker (α-SMA) at the invasive border of lung carcinoma. The positive rates of SNAIL and ZEB1 were 26.7% and 0% in the tumor center and 40% and 20% in tumor margin, respectively. In addition, the expression of both SNAIL and ZEB1 at the border of tumor was observed in two cases (2/10). These two cases were associated with lymph node metastasis and advanced stage. The process of EMT has been suggested to be of prime importance for tissue and lymphovascular invasion. The process of EMT may be activated in the tumor border of lung adenocarcinoma. Related transcription factors, such as SNAIL and SLUG, and ZEB1, might be induced by paracrine effects of surrounded inflammatory cells and fibroblasts.
International journal of clinical and experimental pathology 08/2014; 7(7):4095-103. · 1.89 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Melittin is a cationic, hemolytic peptide that is the main toxic component in the venom of the honey bee (Apis mellifera). It has been used in treatment of various chronic inflammatory diseases. However, the cellular mechanism and the anti-apoptotic effect of melittin in Propionibactierium acnes (P. acnes)-induced THP-1 cells have not been explored. In the present study, we investigated the anti-inflammatory and anti-apoptotic mechanism by examining the effect of melittin on P. acnes-induced THP-1 monocytic cells. THP-1 monocytic cells were stimulated by heat-killed P. acnes in the presence of melittin. The expression levels of pro-inflammatory cytokines, NF-κB signaling, caspase family, and PARP signaling were measured by ELISA or Western blot analysis. The number of apoptotic cells and changes of cell morphology were examined using fluorescence microscopy and flow cytometry. Heat-killed P. acnes increased the secretion of pro-inflammatory cytokines and cleavage of caspase-3 and -8 in heat-killed P. acnes-induced THP-1 cells. However, treatment with melittin inhibited the pro-inflammatory cytokines and cleavage of the caspase-3 and -8. Moreover, the cleaved PARP appeared after 8h of heat-killed P. acnes treatment and its cleavage was reduced by melittin treatment. These results demonstrate that 1.0×10(7) CFU/ml of heat-killed P. acnes-induces THP-1 cell apoptosis and secretion of inflammatory cytokines. Also, administration of melittin significantly decreases the expression of various inflammatory cytokines in heat-killed P. acnes-treated THP-1 monocytic cells. In particular, melittin exerts anti-apoptotic effects against 1.0×10(7) CFU/ml of heat-killed P. acnes injury to THP-1 cells.
European Journal of Pharmacology 07/2014; 740. DOI:10.1016/j.ejphar.2014.06.058 · 2.53 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Epithelial-mesenchymal transition (EMT) has been implicated in embryonic development, fibrosis, and tumor metastasis. Nuclear factor (NF)-κB plays a central role in coordinating the inflammatory and wound healing responses during liver fibrogenesis. However, the involvement of NF-κB during EMT in liver cells remains unidentified. To develop a therapeutic approach for EMT during liver fibrogenesis, we examined the inhibition of transcription factor NF-κB using the decoy oligodeoxynucleotides (ODN) strategy in liver fibrosis in vitro and in vivo. NF-κB decoy ODN contains consensus binding sequences of NF-κB binding site. NF-κB decoy ODN effectively suppresses transforming growth factor (TGF)-β1-induced EMT in AML-12 murine hepatocytes. Liver fibrosis induced by CCl4 administration was suppressed by NF-κB decoy ODN. Furthermore, NF-κB decoy ODN was shown to inhibit the EMT process in fibrotic liver in vivo. This study demonstrates the feasibility of NF-κB decoy ODN treatment for preventing liver fibrosis via EMT processes.
Human Gene Therapy 06/2014; 25(8). DOI:10.1089/hum.2013.106 · 3.76 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Apamin is an integral part of bee venom, as a peptide component. It has long been known as a highly selective block Ca(2+)-activated K(+) (SK) channels. However, the cellular mechanism and anti-fibrotic effect of apamin in TGF-β1-induced hepatocytes have not been explored. In the present study, we investigated the anti-fibrosis or anti-EMT mechanism by examining the effect of apamin on TGF-β1-induced hepatocytes. AML12 cells were seeded at ∼60% confluence in complete growth medium. Twenty-four h later, the cells were changed to serum free medium containing the indicated concentrations of apamin. After 30 min, the cells were treated with 2 ng/ml of TGF-β1 and co-cultured for 48 h. Also, we investigated the effects of apamin on the CCl4-induced liver fibrosis animal model. Treatment of AML12 cells with 2 ng/ml of TGF-β1 resulted in loss of E-cadherin protein at the cell-cell junctions and concomitant increased expression of vimentin. In addition, phosphorylation levels of ERK1/2, Akt, Smad2/3 and Smad4 were increased by TGF-β1 stimulation. However, cells treated concurrently with TGF-β1 and apamin retained high levels of localized expression of E-cadherin and showed no increase in vimentin. Specifically, treatment with 2 μg/ml of apamin almost completely blocked the phosphorylation of ERK1/2, Akt, Smad2/3 and Smad4 in AML12 cells. In addition, apamin exhibited prevention of pathological changes in the CCl4-injected animal models. These results demonstrate the potential of apamin for the prevention of EMT progression induced by TGF-β1 in vitro and CCl4-injected in vivo.
Biochemical and Biophysical Research Communications 05/2014; 450(1). DOI:10.1016/j.bbrc.2014.05.089 · 2.30 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Liver injuries are repaired by fibrosis and regeneration. The cause of fibrosis and diminished regeneration, especially in liver cirrhosis, is still unknown. Epithelial-mesenchymal transition (EMT) has been found to be associated with liver fibrosis. The possibility that EMT could contribute to hepatic fibrogenesis reinforced the concept that activated hepatic stellate cells are not the only key players in the hepatic fibrogenic process and that other cell types, either hepatic or bone marrow-derived cells could contribute to this process. Following an initial enthusiasm for the discovery of this novel pathway in fibrogenesis, more recent research has started to cast serious doubts upon the real relevance of this phenomenon in human fibrogenetic disorders. The debate on the authenticity of EMT or on its contribution to the fibrogenic process has become very animated. The overall result is a general confusion on the meaning and on the definition of several key aspects. The aim of this article is to describe how EMT participates to hepatic fibrosis and discuss the evidence of supporting this possibility in order to reach reasonable and useful conclusions.
World Journal of Hepatology 04/2014; 6(4):207-216. DOI:10.4254/wjh.v6.i4.207
[Show abstract][Hide abstract] ABSTRACT: Melittin is the main component in the venom of the honey bee (Apis mellifera). It has multiple effects including anti-bacterial, anti-viral, and anti-inflammatory activities, in various cell types. However, the anti-inflammatory mechanisms of melittin have not been elucidated in Propionibactierium acnes (P. acnes) induced keratinocyte or inflammatory skin disease animal models. In this study, we examined the effects of melittin on the production of inflammatory cytokines in heat-killed P. acnes-induced HaCaT cells. Heat-killed P. acnes treated keratinocytes increased the expression of pro-inflammatory cytokines and toll like receptor 2. However, melittin treatment significantly suppressed the expression of these cytokines through regulation of the NF-κB and MAPK signaling pathways. Subsequently, we examined the living P. acnes (1 × 10(7) CFU) were intradermally injected into the ear of mice. Living P. acnes injected ears showed cutaneous erythema, swelling, and granulomatous response at 24 h after injection. However, melittin-treated ears showed markedly reduced swelling and granulomatous responses compared with ears injected with only living P. acnes. These results demonstrate the feasibility of applying melittin for the prevention of inflammatory skin diseases induced by P. acnes.Journal of Investigative Dermatology accepted article preview online, 4 February 2014; doi:10.1038/jid.2014.75.
[Show abstract][Hide abstract] ABSTRACT: Honeybee (Apis mellifera L.) venom (BV) has been traditionally used for the treatment of pain and inflammatory diseases such as itchy skin problems. However, the precise mechanism of BV in ameliorating the scratching behavior is not fully understood.
In order to evaluate the effect of BV on atopic dermatitis-related symptoms in mice, we used a mouse skin scratching model induced by compound 48/80. The anti-itch effect of BV was investigated in a compound 48/80-induced mouse scratching behavior model. BALB/c mice were injected intraperitoneally with vehicle (saline 0.9%) or BV (0.01 and 0.1 mg/kg). One hour after treatment, the animals received a subcutaneous injection of compound 48/80. Intraperitoneal administration of BV (0.01 and 0.1 mg/kg) attenuated compound 48/80-induced scratching behaviors. The anti-scratching behavior effect of BV was in proportional to its vascular permeability effects. Treatment with BV also inhibited the degranulation of mast cells and the production of pro-inflammatory cytokines in compound 48/80-treated skin tissues. According to these results, BV may improve atopic dermatitis-related symptoms by inhibiting the mast cell degranulation and pro-inflammatory cytokine expression.
International journal of clinical and experimental pathology 12/2013; 6(12):2896-903. · 1.89 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Renal tubulointerstitial fibrosis is considered to be a common final pathway related to the progressive loss of renal function in chronic kidney disease. It is characterized by the excessive accumulation of extracellular matrix through the pivotal role of epithelial-mesenchymal transition. Transforming growth factor-β1 is postulated to play a central role in renal fibrosis via a downstream pathway such as Smad. Specificity protein 1 (Sp1), which is another transcription factor, is also involved in the basal expression of extracellular matrix. In this study, we investigate the effect of Smad decoy oligodeoxynucleotides (ODN) and Sp1 decoy ODN in unilateral ureteral obstruction induced renal fibrosis in mice. Furthermore, the effectiveness of the newly designed chimeric decoy ODN, which contains both Smad and Sp1 binding sequences in one decoy molecule (Smad/Sp1 chi decoy ODN), was demonstrated. The expression of fibrosis and inflammatory related cytokines and products of fibrosis were ameliorated in the Smad, Sp1 and chimeric decoy ODN treated groups compared with the scrambled decoy ODN treated group. Epithelial-mesenchymal transition was suppressed by the Smad, Sp1 and Smad/Sp1 chi decoy ODN. Immunohistochemistry and Western-blot analysis revealed that Smad/Sp1 chi decoy ODN showed a more significant inhibitory effect on fibrosis and EMT compared with Smad and Sp1 decoy ODNs. These results support the efficacy of Smad/Sp1 chi decoy compared with a single Smad or Sp1 decoy ODNs in preventing renal fibrosis induced by unilateral ureteral obstruction.
[Show abstract][Hide abstract] ABSTRACT: The pathophysiology of chronic renal disease is characterized by a progressive loss of renal function and deposition of the extracellular matrix, leading to widespread tissue fibrosis. Much of the matrix in chronic renal disease is synthesized by interstitial myofibroblasts, recruited from resident fibroblasts and circulating precursors. These changes are believed to be derived from epithelial-mesenchymal transition (EMT) of tubuloepithelial cells. To develop a novel therapeutic approach for treating renal fibrosis, we examined the simultaneous inhibition of the transcription factors NF-κB and Sp1 in a mouse model of unilateral ureteral obstruction (UUO). To simultaneously inhibit both NF-κB and Sp1, we developed chimeric (Chi) decoy oligodeoxynucleotide (ODN) which contained binding sequences for both NF-κB and Sp1 in a single decoy molecule to enhance the effective use of decoy ODN strategy. Chi decoy ODN significantly attenuated tubulointerstitial fibrosis in a mouse model of UUO compared to scrambled decoy ODN, as demonstrated by the reduced interstitial volume, macrophage infiltration, and fibrosis-related gene expression. Interestingly, Chi decoy ODN also regulated EMT-related gene expression, leading to the inhibition of renal fibrotic changes in vivo and in vitro. The present study demonstrates the feasibility of Chi decoy ODN treatment for preventing renal fibrosis and EMT processes. This strategy might be useful to improve the clinical outcome after chronic renal disease.
Journal of Molecular Medicine 11/2012; 91(5). DOI:10.1007/s00109-012-0972-2 · 5.11 Impact Factor