Lactobacillus plantarum LMT1-48 exerts anti-obesity effect in high-fat diet-induced obese mice by regulating expression of lipogenic genes.

Obesity is a major health problem and is known to be closely associated with metabolic diseases. Abnormal hepatic accumulation of fat causes fatty liver or hepatic steatosis, and long-term consumption of a high-fat diet is known to be a key obesity-causing factor. Recent studies have demonstrated that probiotics such as Lactobacillus strains, exert an anti-obesity effect by regulating adipogenesis. However, it is still unknown how the consumption of probiotics can reduce abdominal fat volume by regulating the hepatic expression of lipogenic genes. Therefore, we evaluated the effect of long-term ingestion of L. plantarum LMT1-48 on the expression of lipogenic genes in high-fat diet (HFD)-fed mice. We observed that treatment of 3T3-L1 adipocytes with L. plantarum LMT1-48 extract inhibited their differentiation and lipid accumulation by downregulating lipogenic genes, namely, PPARγ, C/EBPα, FAS, and FABP4. Interestingly, administration of L. plantarum LMT1-48 reduced liver weight and liver triglycerides concurrently with the downregulation of the lipogenic genes PPARγ, HSL, SCD-1, and FAT/CD36 in the liver, resulting in the reduction of body weight and fat volume in HFD-fed obese mice. Notably, we also observed that the administration of at least 106 CFU of L. plantarum LMT1-48 significantly lowered body weight and abdominal fat volume in modified diet-fed mouse models. Collectively, these data suggest that L. plantarum LMT1-48 is a potential healthy food for obese people.

VEGF receptor binding peptide-linked amphiphilic peptide with arginines and valines for endothelial cell-specific gene delivery.

An amphiphilic peptide with a 3-arginine stretch and a 6-valine stretch (R3V6) has been previously reported to deliver plasmid DNA (pDNA) into cells with no toxicity. Here, the vascular endothelial growth factor receptor binding peptide (VRBP) was linked to R3V6 to promote endothelial-specific gene delivery. The pDNA/VRBP-linked R3V6 (VRBP-R3V6) complex was physically characterized via various methods. In a gel retardation assay, pDNA was completely retarded by VRBP-R3V6 at a weight ratio of 1:2 (pDNA:peptide). VRBP-R3V6 also protected pDNA from DNase I for longer than 60 min. Heparin competition assay showed that the pDNA/VRBP-R3V6 complex did not release pDNA when heparin was introduced at a two-fold weight excess of pDNA. In vitro transfection showed that VRBP-R3V6 had transfection efficiency into endothelial cells approximately 200 times greater than that of R3V6. In addition, the transfection efficiency was further enhanced into hypoxic endothelial cells. However, in human embryonic kidney 293 and neuroblastoma N2A cells, VRBP-R3V6 only achieved a transfection rate 10 times higher than R3V6, indicating that VRBP-R3V6 has high specificity for endothelial cells. VRBP-R3V6 was also shown to be nontoxic in a cytotoxicity assay. The data presented here suggest that VRBP-R3V6 may prove useful for specific gene delivery to endothelial cells.

Amphiphilic peptides with arginine and valine residues as siRNA carriers.

An efficient and safe delivery carrier is required for the therapeutic application of siRNA. In this research, amphiphilic peptides with arginine and valine residues were evaluated as siRNA carriers. The peptides were composed of 1-4 arginine-blocks and 6 valine-blocks. In the aqueous solution, the arginine-valine peptides (RV peptides) formed micelles with hydrophobic cores comprised of a valine block and a cationic surface comprised of an arginine block. In a gel retardation assay, the RV peptides completely retarded siRNA at a 1:10 weight ratio (siRNA:peptide). A heparin competition assay suggested that the RV peptides formed more stable complexes with siRNA than they did with polyethylenimine (25 kDa, PEI25k). In an in vitro silencing assay, a dual luciferase expression (Renilla and firefly luciferases) vector, psiCHECK2, was co-transfected into human embryonic kidney 293 cells with Renilla-siRNA using the RV peptides. The specific silencing effect of Renilla luciferase was analyzed in reference to firefly luciferase. The results showed that the R3V6 peptide was more efficient than the R1V6, R2V6, and R4V6 peptides in silencing Renilla luciferase. In the flow cytometry and in vitro silencing studies, the R3V6 peptide delivered Renila-siRNA as efficiently as PEI25k. The siVEGF/R3V6 peptide also reduced endogenous vascular endothelial growth factor (VEGF) expression in CT27 cells as efficiently as PEI25k. A cytotoxicity assay showed that RV peptides did not cause any cytotoxicity. Therefore, RV peptides may be useful for the development of a safe and efficient delivery carrier of siRNA.

Amphiphilic peptides with arginines and valines for the delivery of plasmid DNA.

A non-toxic and efficient gene carrier is one requirement for clinical gene therapy. In this study, amphiphilic peptides composed of arginines and valines were synthesized and characterized as plasmid DNA (pDNA) carriers. The peptides have a cationic region containing 1-4 arginines and a hydrophobic region containing 6 valines. The arginine-valine peptides (RV peptides) formed micelles in aqueous solution with a critical micelle concentration (CMC) of 1.35 mg/ml. In gel retardation assay, the RV peptides retarded all pDNA at weight ratios (pDNA:RV peptide) of 1:3 for R1V6, 1:2 for R2V6 and R3V6, and 1:1 for R4V6. A heparin competition assay showed that the R3V6 peptide formed tighter complexes with pDNA than poly-L-lysine (PLL). In vitro transfection assay into HEK293 cells showed that the R1V6 and R2V6 peptides had the highest transfection efficiencies at 1:30 weight ratios (pDNA:RV peptide), while the R3V6 and R4V6 peptides had the highest efficiencies at 1:20 weight ratios. Under optimal conditions, the R3V6 peptide had the highest transfection efficiency of all the RV peptides and PLL. MTT assay showed that the RV peptides did not have any detectable toxicity to cells. Therefore, the RV peptide may be useful for the development of non-toxic gene carriers.

A case of hypocalcemia-induced dilated cardiomyopathy.

Hypocalcemia is a rare cause of dilated cardiomyopathy. Hypocalcemia induced cardiomyopathy is usually reversible when calcium level returns to normal range. We experienced a case of 57-year-old woman who had suffered from hypocalcemia after total thyroidectomy and its consequence of dilated cardiomyopathy. After supplementation of calcium and vitamin D, symptoms of heart failure and LV function were recovered.