Increased effects of ginsenosides on the expression of cholesterol 7α-hydroxylase but not the bile salt export pump are involved in cholesterol metabolism

An extract from red ginseng [steamed and dried roots of Panax ginseng C.A. Meyer (RGE)] has been shown to have various actions on physiological functions. The mechanisms by which RGE promotes cholesterol metabolism in the liver are unclear, but RGE decreases the plasma levels of cholesterol. We investigated whether RGE affected the mRNA expression of cholesterol metabolism-related proteins such as cytochrome P450 (CYP)7A1 and bile salt export pump (BSEP) in the liver in hypercholesterolemic rats and rat primary hepatocytes. In-vivo studies showed the upregulation of CYP7A1 mRNA in hypercholesterolemic rats treated with RGE. Treatment with RGE exhibited decreased ratios of low-density lipoprotein-cholesterol to high-density lipoprotein-cholesterol compared with hypercholesterolemia without RGE. In-vitro studies also showed the upregulation of CYP7A1 mRNA and protein levels by the addition of RGE to rat primary hepatocytes. The mRNA levels of BSEP exhibited few changes. The sustained levels of the liver X receptor (LXR) in vivo and the increased levels of LXR in vitro on RGE treatment could be involved in the upregulation of CYP7A1. To clarify the effects of 11 ginsenosides including RGE on the mRNA levels of CYP7A1 and BSEP, we performed in-vitro experiments using rat primary hepatocytes. The ginsenosides Ro, Rg₃, Re, Rg₁, and Rg₂ exhibited increased mRNA levels of CYP7A1. These results suggest that several ginsenosides including RGE promoted cholesterol metabolism due to upregulation of CYP7A1.     

Blocking acid-sensing ion channel 1 alleviates Huntington's disease pathology via an ubiquitin-proteasome system-dependent mechanism

Huntington's disease (HD) is a fatal neurodegenerative disorder. Despite a tremendous effort to develop therapeutic tools in several HD models, there is no effective cure at present. Acidosis has been observed previously in cellular and in in vivo models as well as in the brains of HD patients. Here we challenged HD models with amiloride (Ami) derivative benzamil (Ben), a chemical agent used to rescue acid-sensing ion channel (ASIC)-dependent acidotoxicity, to examine whether chronic acidosis is an important part of the HD pathomechanism and whether these drugs could be used as novel therapeutic agents. Ben markedly reduced the huntingtin-polyglutamine (htt-polyQ) aggregation in an inducible cellular system, and the therapeutic value of Ben was successfully recapitulated in the R6/2 animal model of HD. To reveal the mechanism of action, Ben was found to be able to alleviate the inhibition of the ubiquitin-proteasome system (UPS) activity, resulting in enhanced degradation of soluble htt-polyQ specifically in its pathological range. More importantly, we were able to demonstrate that blocking the expression of a specific isoform of ASIC (asic1a), one of the many molecular targets of Ben, led to an enhancement of UPS activity and this blockade also decreased htt-polyQ aggregation in the striatum of R6/2 mice. In conclusion, we believe that chemical compounds that target ASIC1a or pharmacological alleviation of UPS inhibition would be an effective and promising approach to combat HD and other polyQ-related disorders.     

The bite-raised condition in aged SAMP8 mice reduces the expression of glucocorticoid receptors in the dorsal and ventral hippocampus

In the present study, we examined whether the effects induced by the bite-raised condition on glucocorticoid receptor (GR) expression differ between the dorsal and ventral hippocampus in SAMP8 mice. In the bite-raised condition, the number of GR-immunoreactive cells was significantly decreased in both the dorsal and ventral CA1 and dentate gyrus (DG) subfields of the hippocampus compared to control mice, as revealed by immunohistochemical analysis. The decrease in the number of GR-immunoreactive cells tended to be greater in the dorsal hippocampus than in the ventral hippocampus. Only in the DG subfield was there a significant difference in the number of GR-immunoreactive cells between the dorsal and ventral hippocampus. These findings suggest that in aged SAMP8 mice, the bite-raised condition decreases the number of GR-immunoreactive cells in both the dorsal and ventral hippocampus.     

Expression and Roles of MMP-2, MMP-9, MMP-13, TIMP-1, and TIMP-2 in Allergic Nasal Mucosa

Purpose

Allergic rhinitis (AR) and asthma share many characteristics, but structural changes are observed far less often in AR. Matrix metalloproteinases (MMPs) constitute a family of Zn-dependent endopeptidases that can decompose the extracellular matrix and basement membrane, and regulate cell infiltration. We analyzed the expression of MMPs and their inhibitors, tissue inhibitors of metalloproteinases (TIMPs), in allergic nasal mucosa after nasal allergen challenge (NAC) and determined their relationship to inflammatory cells.

Methods

Nasal mucosa specimens were obtained at surgery performed for hypertrophied turbinates. We performed NAC with house dust mite (HDM) allergen disks and control disks, and took biopsies at 30 minutes, 6 hours, and 12 hours after NAC. Cells expressing MMP-2, MMP-9, MMP-13, TIMP-1, and TIMP-2, as well as eosinophils and mast cells, were analyzed immunohistochemically. The MMPs and TIMPs in allergic nasal mucosa were quantified using enzyme-linked immunosorbent assays.

Results

At 30 minutes post-NAC, HDM-exposed nasal mucosa exhibited significantly more MMP-2+, MMP-9+, MMP-13+, TIMP-1+, and TIMP-2+ cells compared with control mucosa, and the numbers of MMP-9+ and TIMP-1+ cells correlated strongly with the number of mast cells. At 6 hours post-NAC, the numbers of MMP+ and TIMP+ cells did not differ significantly between HDM-exposed mucosa and control mucosa, but the ratios of MMP+ cells to TIMP+ cells were higher in HDM-exposed mucosa. At 12 hours post-NAC, the number of MMP-13+ cells tended to be higher in HDM-exposed mucosa and was strongly correlated with the number of eosinophils. Quantitatively, the levels of MMP-2 and MMP-13 were significantly higher than the MMP-9 level, and the TIMP-2 level was significantly higher than the TIMP-1 level in allergic nasal mucosa.

Conclusions

We demonstrated increased expression of MMP-2, MMP-9, and MMP-13 in allergic nasal mucosa, high MMPs-to-TIMP-1 ratios, and a strong correlation between MMP-9 and mast cells and between MMP-13 and eosinophils. The imbalance between MMPs and TIMPs may contribute to the migration of inflammatory cells such as eosinophils and mast cells to the nasal mucosa of AR patients, suggesting a possible active role of MMPs in AR.     

Utility of immunohistochemical analysis of KAI1, epithelial-specific antigen, and epithelial-related antigen for distinction of chromophobe renal cell carcinoma, an eosinophilic variant from renal oncocytoma

Distinction of renal oncocytoma (RO) from chromophobe renal cell carcinoma (ChRCC) is important because their clinical behavior is different. As part of a search for the best available immunohistochemical markers to distinguish ChRCC from RO, we investigated the immunohistochemical profiles of these tumors. We selected 30 renal tumors consisting of ChRCC, typical variant (n = 14), ChRCC, eosinophilic variant (n = 6), and RO (n = 10). Their expression of cytokeratin (CK) 7, KAI1, epithelial-specific antigen (ESA), epithelial-related antigen (ERA), Claudin- 7, and Claudin-8 was studied using an autostainer. Immunoreactivity was assessed based on a combined score of the extent and intensity of staining. Compared to RO, a significantly higher percentage of the total ChRCCs stained positive for CK7 (85% vs. 10%, respectively), KAI1 (90% vs. 10%), ESA (95% vs. 10%), ERA (95% vs. 10%), and Claudin-7 (95% vs. 20%) (P < 0.001). Additionally, there was a significant difference between the percentage of ChRCC eosinophilic variant (ChRCC-E) and RO that stained positive for KAI1 (100% vs. 10%, respectively), ESA (83% vs. 10%), and ERA (83% vs. 10%) (P < 0.001). We recommend immunohistochemical analysis of KAI1, ESA, and ERA to distinguish ChRCC-E from RO.     

Low-carbohydrate diet by staple change attenuates postprandial GIP and CPR levels in type 2 diabetes patients

AimsThe aim of this study is to investigate the effects of a low-carbohydrate staple food (i.e., low-carbohydrate bread) on glucose and lipid metabolism and pancreatic and enteroendocrine hormone secretion in comparison with meals containing normal-carbohydrate bread, without consideration of the carbohydrate content of the side dishes.MethodsT2DM patients (n = 41) were provided meals containing low-carbohydrate bread (LB) together with side dishes or normal-carbohydrate bread (NB) together with side dishes every other day as a breakfast. Blood glucose levels were evaluated by using a continuous glucose monitoring system; blood samples were collected before and 1 and 2 h after the breakfast.ResultsPostprandial blood glucose levels, plasma insulin, plasma glucose-dependent insulinotropic polypeptide (GIP) and plasma triglyceride were significantly lower and plasma glucagon levels were significantly higher in LB compared with those in NB. Plasma glucagon-like peptide-1 (GLP-1) levels did not differ in the LB and NB groups.ConclusionsThese results indicate that changing only the carbohydrate content of the staple food has benefits on glucose and lipid metabolism in T2DM patients concomitant with the decrease of insulin and GIP secretion, which ameliorate body weight gain and insulin resistance.     

Single chain variable fragment antibodies against Shiga toxins isolated from a human antibody phage display library

Shiga toxins (Stxs) are involved in the pathogenesis of hemolytic-uremic syndrome and other severe systemic complications following enterohemorrhagic Escherichia coli infection in humans. Passive immunotherapies using monoclonal antibodies have been shown to be effective for neutralizing the toxic effects of Stxs. However, animal-derived monoclonal antibodies are sometimes immunogenic and their production is both laborious and expensive. We here report the isolation of single-chain variable fragment antibodies against Stxs by screening a phage display library constructed from a naïve human repertoire. An antibody among the selected clones designated B22 bound to the binding subunits of both Stx-1 and Stx-2, and strongly neutralized the cytotoxicity of Stx-1. This is the first example of a monovalent antibody showing Stx-neutralizing activity. The B22 antibody is also completely naturally occurring in human, which reduces the possibility of adverse immunological effects, and can be easily produced using bacterial protein synthesis systems.     

Inhibition of Atherosclerotic Plaque Development by Oral Administration of α-Glucosyl Hesperidin and Water-Dispersible Hesperetin in Apolipoprotein E Knockout Mice

Objective: Hesperidin, an abundant flavonoid in citrus fruit, and its aglycone, hesperetin, have been reported to possess various physiological activities, including antioxidant, anti-inflammatory, hypolipidemic, and antihypertensive activities. In this study, we investigated whether α-glucosyl hesperidin and water-dispersible hesperetin have protective effects on atherosclerotic progression in apolipoprotein E knockout (Apo-E KO) mice.

Methods: Ten-week-old male Apo-E KO mice were randomly assigned a regular high-fat diet, a high-fat diet with 0.5% α-glucosyl hesperidin, or a high-fat diet with 0.1% water-dispersible hesperetin for 12?weeks. Measurement of plasma total cholesterol levels, histological staining of aortic root, and immunohistochemistry for macrophages were performed to evaluate atherosclerotic plaque formation. Vascular reactivity of mouse aortic rings was also measured.

Results: Both α-glucosyl hesperidin and water-dispersible hesperetin reduced plasma total cholesterol level. They also reduced plaque formation area, adipose deposition, and macrophage infiltration into atherosclerotic lesion. Vascular-endothelium-dependent relaxation in response to acetylcholine was improved in both experimental diet groups compared to the high-fat diet group.

Conclusions: Our study suggests that both α-glucosyl hesperidin and water-dispersible hesperetin exert protective effects on atherosclerotic progression in Apo-E KO mice because they exhibit hypolipidemic activity, reduce inflammation through macrophages, and prevent endothelial dysfunction.