ROCK1 regulates insulin secretion from ß-cells.

OBJECTIVE: The endocrine pancreatic ß-cells play a pivotal role in maintaining whole-body glucose homeostasis and its dysregulation is a consistent feature in all forms of diabetes. However, knowledge of intracellular regulators that modulate ß-cell function remains incomplete. We investigated the physiological role of ROCK1 in the regulation of insulin secretion and glucose homeostasis. METHODS: Mice lacking ROCK1 in pancreatic ß-cells (RIP-Cre; ROCK1loxP/loxP, ß-ROCK1-/-) were studied. Glucose and insulin tolerance tests as well as glucose-stimulated insulin secretion (GSIS) were measured. An insulin secretion response to a direct glucose or pyruvate or pyruvate kinase (PK) activator stimulation in isolated islets from ß-ROCK1-/- mice or ß-cell lines with knockdown of ROCK1 was also evaluated. A proximity ligation assay was performed to determine the physical interactions between PK and ROCK1. RESULTS: Mice with a deficiency of ROCK1 in pancreatic ß-cells exhibited significantly increased blood glucose levels and reduced serum insulin without changes in body weight. Interestingly, ß-ROCK1-/- mice displayed a progressive impairment of glucose tolerance while maintaining insulin sensitivity mostly due to impaired GSIS. Consistently, GSIS markedly decreased in ROCK1-deficient islets and ROCK1 knockdown INS-1 cells. Concurrently, ROCK1 blockade led to a significant decrease in intracellular calcium and ATP levels and oxygen consumption rates in isolated islets and INS-1 cells. Treatment of ROCK1-deficient islets or ROCK1 knockdown ß-cells either with pyruvate or a PK activator rescued the impaired GSIS. Mechanistically, we observed that glucose stimulation in ß-cells greatly enhanced ROCK1 binding to PK. CONCLUSIONS: Our findings demonstrate that ß-cell ROCK1 is essential for glucose-stimulated insulin secretion and for glucose homeostasis and that ROCK1 acts as an upstream regulator of glycolytic pyruvate kinase signaling.

Control of type O foot-and-mouth disease by vaccination in Korea, 2014-2015.

On December 3, 2014, a type O foot-and-mouth disease (FMD) outbreak began in Korea. Although vaccinations were administered, FMD cases increased steadily for five months, and reached 185 cases by April 2015. Most of the affected animals were pigs, which are vulnerable to vaccination. The FMD virus belonged to the South-East Asia (SEA) topotype that had been observed three times in Korea between April 2010 and July 2014. However, the FMD virus isolated in December 2014 had a unique feature; that is, partial deletion of the 5´ non-coding region, a deletion not seen in previous SEA topotype isolates identified in Korea. We conclude that this outbreak included the introduction of a new FMD strain to Korea, and that Korea was now affected by genetically similar FMD virus strains that are related to those from neighboring countries.

Characterization of Echinostoma cinetorchis endoribonuclease, RNase H.

Echinostoma cinetorchis is an oriental intestinal fluke causing significant pathological damage to the small intestine. The aim of this study was to determine a full-length cDNA sequence of E. cinetorchis endoribonuclease (RNase H; EcRNH) and to elucidate its molecular biological characters. EcRNH consisted of 308 amino acids and showed low similarity to endoribonucleases of other parasites (<40%). EcRNH had an active site centered on a putative DDEED motif instead of DEDD conserved in other species. A recombinant EcRNH produced as a soluble form in Escherichia coli showed enzymatic activity to cleave the 3'-O-P bond of RNA in a DNA-RNA duplex, producing 3'-hydroxyl and 5'-phosphate. These findings may contribute to develop antisense oligonucleotides which could damage echinostomes and other flukes.

ACE insertion/deletion polymorphism is associated with periodontal disease in Korean population.

OBJECTIVE: Angiotensin-converting enzyme (ACE) is the core enzyme in the renin-angiotensin system (RAS), which catalyzes the production of angiotensin II (Ang II). The aim of this study was to determine whether ACE gene is associated with the development of the periodontal disease. DESIGN: To investigate whether ACE is involved in the development of the periodontal disease, 199 periodontal disease patients and 165 control subjects were studied. The ACE insertion/deletion polymorphism was analyzed using polymerase chain reaction (PCR). SNPStats and SPSS 18.0 were used for the analysis of genetic data. Logistic regression models were performed to determine odds ratio (OR), 95% confidence interval (CI), and P value. RESULTS: Genotypic frequencies of I/I, I/D, and D/D were 25.4%, 42.3%, and 32.3% vs. 35.3%, 41.7%, and 23.1% (periodontal disease group vs. control group), respectively. In the genotype analysis of the ACE insertion/deletion polymorphism, codominant and log-additive models both showed significant association with periodontal disease [OR = 1.94, 95% CI = 1.05-3.61, P=0.036 in the codominant model (I/I vs. D/D); OR = 1.39, 95% CI = 1.02-1.90, P = 0.034 in the log-additive model (I/I vs. I/D vs. D/D)]. CONCLUSIONS: These results suggest that the ACE insertion/deletion polymorphism may be associated with the susceptibility to the periodontal disease in the Korean population.

Development of a polymerase chain reaction applicable to rapid and sensitive detection of Clonorchis sinensis eggs in human stool samples.

Microscopic examination of eggs of parasitic helminths in stool samples has been the most widely used classical diagnostic method for infections, but tiny and low numbers of eggs in stool samples often hamper diagnosis of helminthic infections with classical microscopic examination. Moreover, it is also difficult to differentiate parasite eggs by the classical method, if they have similar morphological characteristics. In this study, we developed a rapid and sensitive polymerase chain reaction (PCR)-based molecular diagnostic method for detection of Clonorchis sinensis eggs in stool samples. Nine primers were designed based on the long-terminal repeat (LTR) of C. sinensis retrotransposon1 (CsRn1) gene, and seven PCR primer sets were paired. Polymerase chain reaction with each primer pair produced specific amplicons for C. sinensis, but not for other trematodes including Metagonimus yokogawai and Paragonimus westermani. Particularly, three primer sets were able to detect 10 C. sinensis eggs and were applicable to amplify specific amplicons from DNA samples purified from stool of C. sinensis-infected patients. This PCR method could be useful for diagnosis of C. sinensis infections in human stool samples with a high level of specificity and sensitivity.

Identification and characterization of a mitochondrial manganese superoxide dismutase of Spirometra erinacei.

A gene encoding the manganese superoxide dismutase (Mn-SOD) of Spirometra erinacei was identified, and the biochemical properties of the recombinant enzyme were partially characterized. The S. erinacei Mn-SOD gene consisted of 669 bp, which encoded 222 amino acids. A sequence analysis of the gene showed that it had typical molecular structures, including characteristic metal-binding residues and motifs that were conserved in Mn-SODs. An analysis of the N-terminal presequence of S. erinacei Mn-SOD revealed that it had physiochemical characteristics commonly found in mitochondria-targeting sequences and predicted that the enzyme is located in the mitochondria. A biochemical analysis also revealed that the enzyme is a typical Mn-SOD. The enzyme was consistently expressed in both S. erinacei plerocercoid larvae and adult worms. Our results collectively suggested that S. erinacei Mn-SOD is a typical mitochondrial Mn-SOD and may play an important role in parasite physiology, detoxifying excess superoxide radicals generated in the mitochondria.