Aristolochia manshuriensis Kom ethyl acetate extract protects against high-fat diet-induced non-alcoholic steatohepatitis by regulating kinase phosphorylation in mouse

Aristolochia manshuriensis Kom (AMK) is an herb used as a traditional medicine; however, it causes side effects such as nephrotoxicity and carcinogenicity. Nevertheless, AMK can be applied in specific ways medicinally, including via ingestion of low doses for short periods of time. Non-alcoholic steatohepatitis (NASH) induced the hepatocyte injury and inflammation. The protective effects of AMK against NASH are unclear; therefore, in this study, the protective effects of AMK ethyl acetate extract were investigated in a high-fat diet (HFD)-induced NASH model. We found decreased hepatic steatosis and inflammation, as well as increased levels of lipoproteins during AMK extract treatment. We also observed decreased hepatic lipid peroxidation and triglycerides, as well as suppressed hepatic expression of lipogenic genes in extract-treated livers. Treatment with extract decreased the activation of c-jun N-terminal kinase 1/2 (JNK1/2) and increased the phosphorylation of extracellular signal-regulated kinase 1/2 (ERK1/2). These results demonstrate that the protective effect of the extract against HFD-induced NASH occurred via reductions in reactive oxygen species production, inflammation suppression, and apoptosis related to the suppression of JNK1/2 activation and increased ERK1/2 phosphorylation. Taken together, these results indicate that that ethyl acetate extract of AMK has potential therapeutic effects in the HFD-induced NASH mouse model.

Erratum

No abstract available.

Human leukocytes regulate ganglioside expression in cultured micro-pig aortic endothelial cells / 한국실험동물학회지

Gangliosides are ubiquitous components of the membranes of mammalian cells that are thought to play important roles in various cell functions such as cell-cell interaction, cell adhesion, cell differentiation, growth control, and signaling. However, the role that gangliosides play in the immune rejection response after xenotransplantation is not yet clearly understood. In this study, the regulatory effects of human leukocytes on ganglioside expression in primary cultured micro-pig aortic endothelial cells (PAECs) were investigated. To determine the impact of human leukocytes on the expression of gangliosides in PAECs, we performed high-performance thin layer chromatography (HPTLC) in PAECs incubated with FBS, FBS containing human leukocytes, human serum containing human leukocytes, and FBS containing TNF-alpha. Both HPTLC and immunohistochemistry analyses revealed that PAECs incubated with FBS predominantly express the gangliosides GM3, GM1, and GD3. However, the expression of GM1 significantly decreased in PAECs incubated for 5 h with TNF-alpha (10 ng/mL), 10% human serum containing human leukocytes, and 10% FBS containing human leukocytes. Taken together, these results suggest that human leukocytes induced changes in the expression profile of ganglioside GM1 similar to those seen upon treatment of PAECs with TNF-alpha. This finding may be relevant for designing future therapeutic strategies intended to prolong xenograft survival.

Relationship between ganglioside expression and anti-cancer effects of the monoclonal antibody against epithelial cell adhesion molecule in colon cancer

The human colorectal carcinoma-associated GA733 antigen epithelial cell adhesion molecule (EpCAM) was initially described as a cell surface protein selectively expressed in some myeloid cancers. Gangliosides are sialic acid-containing glycosphingolipids involved in inflammation and oncogenesis. We have demonstrated that treatment with anti-EpCAM mAb and RAW264.7 cells significant inhibited the cell growth in SW620 cancer cells, but neither anti-EpCAM mAb nor RAW264.7 cells alone induced cytotoxicity. The relationship between ganglioside expression and the anti-cancer effects of anti-EpCAM mAb and RAW264.7 was investigated by high-performance thin-layer chromatography. The results demonstrated that expression of GM1 and GD1a significantly increased in the ability of anti-EpCAM to inhibit cell growth in SW620 cells. Anti-EpCAM mAb treatment increased the expression of anti-apoptotic proteins such as Bcl-2, but the expression of pro-apoptotic proteins Bax, TNF-alpha, caspase-3, cleaved caspase-3, and cleaved caspase-8 were unaltered. We observed that anti-EpCAM mAb significantly inhibited the growth of colon tumors, as determined by a decrease in tumor volume and weight. The expression of anti-apoptotic protein was inhibited by treatment with anti-EpCAM mAb, whereas the expression of pro-apoptotic proteins was increased. These results suggest that GD1a and GM1 were closely related to anticancer effects of anti-EpCAM mAb. In light of these results, further clinical investigation should be conducted on anti-EpCAM mAb to determine its possible chemopreventive and/or therapeutic efficacy against human colon cancer.

Roles of gangliosides in mouse embryogenesis and embryonic stem cell differentiation

Gangliosides have been suggested to play important roles in various functions such as adhesion, cell differentiation, growth control, and signaling. Mouse follicular development, ovulation, and luteinization during the estrous cycle are regulated by several hormones and cell-cell interactions. In addition, spermatogenesis in seminiferous tubules of adult testes is also regulated by several hormones, including follicle-stimulating hormone (FSH) and luteinizing hormone (LH) and cell-cell interactions. The regulation of these processes by hormones and cell-cell interactions provides evidence for the importance of surface membrane components, including gangliosides. During preimplantation embryo development, a mammalian embryo undergoes a series of cleavage divisions whereby a zygote is converted into a blastocyst that is sufficiently competent to be implanted in the maternal uterus and continue its development. Mouse embryonic stem (mES) cells are pluripotent cells derived from mouse embryo, specifically, from the inner cell mass of blastocysts. Differentiated neuronal cells are derived from mES cells through the formation of embryonic bodies (EBs). EBs recapitulate many aspects of lineage-specific differentiation and temporal and spatial gene expression patterns during early embryogenesis. Previous studies on ganglioside expression during mouse embryonic development (including during in vitro fertilization, ovulation, spermatogenesis, and embryogenesis) reported that gangliosides were expressed in both undifferentiated and differentiated (or differentiating) mES cells. In this review, we summarize some of the advances in our understanding of the functional roles of gangliosides during the stages of mouse embryonic development, including ovulation, spermatogenesis, and embryogenesis, focusing on undifferentiated and differentiated mES cells (neuronal cells).

The roles of glycosphingolipids in the proliferation and neural differentiation of mouse embryonic stem cells

Glycosphingolipids including gangliosides play important regulatory roles in cell proliferation and differentiation. UDP-glucose:ceramide glucosyltransferase (Ugcg) catalyze the initial step in glycosphingolipids biosynthesis pathway. In this study, Ugcg expression was reduced to approximately 80% by short hairpin RNAs (shRNAs) to evaluate the roles of glycosphingolipids in proliferation and neural differentiation of mouse embryonic stem cells (mESCs). HPTLC/immunofluorescence analyses of shRNA-transfected mESCs revealed that treatment with Ugcg-shRNA decreased expression of major gangliosides, GM3 and GD3. Furthermore, MTT and Western blot/immunofluorescence analyses demonstrated that inhibition of the Ugcg expression in mESCs resulted in decrease of cell proliferation (P < 0.05) and decrease of activation of the ERK1/2 (P < 0.05), respectively. To further investigate the role of glycosphingolipids in neural differentiation, the embryoid bodies formed from Ugcg-shRNA transfected mESCs were differentiated into neural cells by treatment with retinoic acid. We found that inhibition of Ugcg expression did not affect embryoid body (EB) differentiation, as judged by morphological comparison and expression of early neural precursor cell marker, nestin, in differentiated EBs. However, RT-PCR/immunofluorescence analyses showed that expression of microtubule- associated protein 2 (MAP-2) for neurons and glial fibrillary acidic protein (GFAP) for glial cells was decreased in neural cells differentiated from the shRNA-transfected mESCs. These results suggest that glycosphingolipids are involved in the proliferation of mESCs through ERK1/2 activation, and that glycosphingolipids play roles in differentiation of neural precursor cells derived from mESCs.

Effects of endocrine disrupting chemicals on expression of phospholipid hydroperoxide glutathione peroxidase mRNA in rat testes

Phospholipid hydroperoxide glutathione peroxidase(PHGPx), an antioxidative selenoprotein, is modulated byestrogen in the testis and oviduct. To examine whetherpotential endocrine disrupting chemicals (EDCs) affectthe microenvironment of the testes, the expression patternsof PHGPx mRNA and histological changes were analyzedin 5-week-old Sprague-Dawley male rats exposed to severalEDCs such as an androgenic compound [testosterone (50,200, and 1,000microg/kg)], anti-androgenic compounds [flutamide(1, 5, and 25mg/kg), ketoconazole (0.2 and 1mg/kg), anddiethylhexyl phthalate (10, 50, and 250mg/kg)], andestrogenic compounds [nonylphenol (10, 50, 100, and 250mg/kg), octylphenol (10, 50, and 250mg/kg), and diethyl-stilbestrol (10, 20, and 40microg/kg)] daily for 3 weeks via oraladministration. Mild proliferation of germ cells andhyperplasia of interstitial cells were observed in the testesof the flutamide-treated group and deletion of thegerminal epithelium and sloughing of germ cells wereobserved in testes of the diethylstilbestrol-treated group.Treatment with testosterone was shown to slightly decreasePHGPx mRNA levels in testes by the reverse transcription-polymerase chain reaction. However, anti-androgeniccompounds (flutamide, ketoconazole, and diethylhexylphthalate) and estrogenic compounds (nonylphenol,octylphenol, and diethylstilbestrol) significantly up-regulated PHGPx mRNA in the testes (p<0.05). Thesefindings indicate that the EDCs might have a detrimentaleffect on spermatogenesis via abnormal enhancement ofPHGPx expression in testes and that PHGPx is useful as abiomarker for toxicity screening of estrogenic or anti-androgenic EDCs in testes.

Dynamic changes of gangliosides expression during the differentiation of embryonic and mesenchymal stem cells into neural cells

Stem cells are used for the investigation of developmental processes at both cellular and organism levels and offer tremendous potentials for clinical applications as an unlimited source for transplantation. Gangliosides, sialic acid-conjugated glycosphingolipids, play important regulatory roles in cell proliferation and differentiation. However, their expression patterns in stem cells and during neuronal differentiation are not known. Here, we investigated expression of gangliosides during the growth of mouse embryonic stem cells (mESCs), mesenchymal stem cells (MSCs) and differentiated neuronal cells by using high-performance thin-layer chromatography (HPTLC). Monosialoganglioside 1 (GM1) was expressed in mESCs and MSCs, while GM3 and GD3 were expressed in embryonic bodies. In the 9-day old differentiated neuronal cells from mESCs cells and MSCs, GM1 and GT1b were expressed. Results from immunostaining were consistent with those observed by HPTLC assay. These suggest that gangliosides are specifically expressed according to differentiation of mESCs and MSCs into neuronal cells and expressional difference of gangliosides may be a useful marker to identify differentiation of mESCs and MSCs into neuronal cells.