Kung-Woo Nam

Bacterial Community Dynamics of Salted and Fermented Shrimp based on Denaturing Gradient Gel Electrophoresis

The Korean traditional seafood jeotgal is consumed directly or as an additive in other foods to improve flavor or fermentation efficiency. Saeujot, made from salted and fermented tiny shrimp (SFS; Acetes japonicus), is the best-selling jeotgal in Korea. In this study, we reveal the microbial diversity and dynamics in naturally fermented shrimp by denaturing gradient gel electrophoresis (DGGE). The population fingerprints of the predominant microbiota and its succession were generated by DGGE analysis of universal V3 16S rDNA polymerase chain reaction (PCR) amplicons. Overall, 17 strains were identified from sequencing of 30 DGGE bands. The DGGE profiles showed diverse bacterial populations in the sample, throughout the fermentation of SFS. Staphylococcus equorum, Halanaerobium saccharolyticum, Salimicrobium luteum, and Halomonas jeotgali were the dominant bacteria, and their levels steadily increased during the fermentation process. Certain other bacteria, such as Psychrobacter jeotgali and Halomonas alimentaria appeared during the early-fermentation process, while Alkalibacterium putridalgicola, Tetragenococcus muriaticus, and Salinicoccus jeotgali appeared during the late-fermentation process. The members of the order Bacillales were found to be predominant during the fermentation of SFS. Furthermore, S. equorum was identified as the dominant bacterial isolate by the traditional method of culturing under aerobic and facultative anaerobic conditions. We expect that this information will facilitate the design of autochthonous starter cultures for the production of SFS with desired characteristic sensory profiles and shorter ripening times.

Ursolic Acid Reduces Mycobacterium tuberculosis- Induced Nitric Oxide Release in Human Alveolar A549 cells

Alveolar epithelial cells have been functionally implicated in Mycobacterium tuberculosis infection. This study investigated the role of ursolic acid (UA)—a triterpenoid carboxylic acid with potent antioxidant, anti-tumor, anti-inflammatory, and anti-tuberculosis properties in mycobacterial infection of alveolar epithelial A549 cells. We observed that M. tuberculosis successfully entered A549 cells. Cytotoxi-city was mediated by nitric oxide (NO). A549 toxicity peaked along with NO generation 72 h after infection. The NO generated by mycobacterial infection in A549 cells was insufficient to kill mycobacteria, as made evident by the mycobacteria growth indicator tube time to detect (MGIT TTD) and viable cell count assays. Treatment of mycobacteria-infected cells with UA reduced the expression of inducible nitric oxide synthase, NO generation, and eventually improved cell viability. Moreover, UA was found to quench the translocation of the transcription factor, nuclear factor kappa B (NF-κB), from the cytosol to the nucleus in mycobacteria-infected cells. This study is the first to demonstrate the cytotoxic role of NO in the eradication of mycobacteria and the role of UA in reducing this cytotoxicity in A549 cells.

In vitro activity of (-)-deoxypergularinine, on its own and in combination with anti-tubercular drugs, against resistant strains of Mycobacterium tuberculosis.

BACKGROUND The increasing incidence of multidrug-resistant tuberculosis (MDR-TB) infections has created a need for new effective drugs that also target extensively drug-resistant tuberculosis (XDR-TB) and/or augment the activities of existing drugs against tuberculosis. AIM This study searched natural products for a new lead compound that targets MDR/XDR-TB. METHODS An active compound was purified from the roots of Cynanchum atratum Bunge (Asclepiadaceae) after screening 1640 plant extracts, and its inhibitory effects against MDR/XDR strains and synergistic effects with existing anti-TB drugs were assessed using the resazurin, MGIT, and checkboard assays. RESULTS (-)-Deoxypergularinine, purified from the roots of C. atratum, inhibited not only M. tuberculosis but also MDR/XDR strains. The minimum inhibitory concentrations (MICs) of (-)-deoxypergularinine for H37Ra, H37Rv, MDR, and XDR strains were all about 12.5 µg/ml. Moreover, combinations of (-)-deoxypergularinine with the first-line standard drugs rifampicin or isoniazid afforded six- and eight-fold reductions in drug MIC values, respectively, against strain H37Ra. CONCLUSIONS (-)-Deoxypergularinine exerts anti-tubercular activities not only against normal tuberculosis strains but also MDR/XDR strains, and synergic effects with rifampicin and isoniazid for the H37Ra strain. The alkaloid may be valuable for targeting M/XDR M. tuberculosis.

Naphthofuroquinone derivatives show strong antimycobacterial activities against drug-resistant Mycobacteria

Tuberculosis, one of the world’s major health problems, has become more serious due to the emergence of multi-drug resistant (MDR) and extensively drug-resistant (XDR) Mycobacterium tuberculosis (MTB). In this study, we performed three anti-MTB assays to evaluate the anti-mycobacterial activity of naphthofuroquinone derivatives against drug-resistant MTB. Among them, methyl 5-[2-(dimethylamino)ethoxy]-7,12-dioxo-7,12-dihydrodinaphtho[1,2-b:2′,3′-d]furan-6-carboxylate (DFC2) exhibited strong anti-mycobacterial activity against MTB H37Ra, H37Rv and four drug-resistant MTB strains. The MIC of DFC2 ranged from 0.19–0.39 μg/ml to 0.78–1.56 μg/ml against all tested MTB strains. Moreover, DFC2 showed low cytotoxicity against fibroblast cells (L929) at concentrations 10–40-fold higher than their MICs. The IC50 value of DFC2 against L929 cells was 15.218 μg/ml. In addition, DFC2 reduced the number of intracellular M. tuberculosis in macrophages in a dose-dependent manner. Taken together, our results indicate DFC2 to be promising new candidate agents for the treatment of tuberculosis.

Pretreatment with recombinant human interleukin 2 (rhIL-2) Up-regulates PCNA-positive cells after partial hepatectomy in rat liver

We prepared recombinant human interleukin-2 (rhIL-2) and studied its pretreated influence on liver regeneration and the blood profile in partially (67%) hepatectomized (PH) male Sprague-Dawley rats. Rats were injected in the tail vein with rhIL-2 three times per day for 3 consecutive days and 67% underwent a partial hepatectomy (PH). Five days after the PH, liver tissue and blood samples were analyzed for liver regeneration and hematological changes. The weight of the liver in the rhIL-2 pretreated groups increased in a dose-dependent manner; with the highest treatment (24 × 104 IU/kg), the maximum liver weight of 88.6% was exhibited. The control group showed a gradual increase to 76.3% of the original liver weight. A histological analysis of the liver showed an increase in proliferating cell nuclear antigen (PCNA)-positive cells in rhIL-2 pretreated rat livers. The rate of hepatocyte proliferation also increased significantly in primary cultured rat liver cells following rhIL-2 treatment. These results suggest that pretreatment with rhIL-2 may play adjuvant roles in liver regeneration after PH.

Administration of rhIL-2 upregulates HGF in the cirrhotic liver of partial hepatectomized rats

Abstract Liver regeneration is a process involving highly organized and ordered tissue growth triggered by the loss of liver tissue. This process has remained a fascinating topic in medicine. In this study, we examined the effects of recombinant human interleukin-2 (rhIL-2) in rats under cirrhotic conditions. Thirty rats were divided into three groups, and liver cirrhosis was induced by injecting diethylnitrosamine for 2 weeks and CCl4 for 8 weeks. After induction of cirrhosis, rhIL-2 was injected daily into their tail vein for three days and then hepatocyte growth factor (HGF), immunocytes in the blood, and TNF-α production were analyzed. The analysis showed that TNF-α was significantly downregulated in the liver without significant changes in nitric oxide and caspase-3/7 activities in primary cultured liver cells. In terms of the number of immunocytes, there were no significant differences between the control and the treated groups. However, HGF levels were significantly higher in the treated groups than the control (saline administration) group. Treatment of rhIL-2 significantly increased the proliferation of primary cultured liver cells. The proliferation rates of primary cultured liver cells were correlated with the upregulation of HGF by treatment with rhIL-2 in vivo and in vitro. These results suggest that rhIL-2 affects liver regeneration, which is at least related to the upregulation of HGF under cirrhotic conditions in rats.

In vitro Antitubercular Activity of 3‐Deoxysappanchalcone Isolated From the Heartwood of Caesalpinia sappan Linn.

Responsible for nearly 1.5 million deaths every year, the infectious disease tuberculosis remains one of the most serious challenges to global health. The emergence of multidrug‐resistant tuberculosis and, more recently, extensively drug‐resistant tuberculosis poses a significant threat in our effort to control this epidemic. New drugs are urgently needed to combat the growing threat of antimicrobial resistance. To achieve this goal, we screened approximately 500 species of medicinal plant methanol extracts and their solvent partitioned fractions for potential inhibitors of Mycobacterium tuberculosis growth. Using microdilution screening, the ethyl acetate solvent partitioned fraction from the heartwood of Caesalpinia sappan exhibited strong antitubercular activity. We isolated the active compound and identified it as 3‐deoxysappanchalcone. The extracted 3‐deoxysappanchalcone possessed activity against both drug‐susceptible and drug‐resistant strains of M. tuberculosis at MIC50s of 3.125–12.5 μg/mL in culture broth and MIC50s of 6.25–12.5 μg/mL inside macrophages and pneumocytes. 3‐Deoxysappanchalcone was also found to act in partial synergy with streptomycin/ethambutol against M. tuberculosis H37Rv. 3‐Deoxysappanchalcone had no cytotoxicity against the A549 cell line up to a concentration of 100 μg/mL (selectivity index > 8–32). Further studies are warranted to establish the in vivo effect and therapeutic potential of 3‐deoxysappanchalcone. Copyright

Polydeoxyribonucleotide administration promotes the expression of proliferating cell nuclear antigen during liver regeneration in rats

Polydeoxyribonucleotide (PDRN) is an agonist of the A2A adenosine receptors. We evaluated the effects of PDRN on liver regeneration induced by partial hepatectomy (PH) in rats, focusing on cell proliferation. Proliferating cell nuclear antigen (PCNA) as an indicator of cell proliferation was targeted to determine whether PDRN treatment could promote proliferation of hepatic cells in regenerating liver. Male Sprague-Dawley rats were divided into three groups: normal group (n = 3) without PH (sham operated), control groups (n = 12) with saline injection after PH, and experimental groups (n = 12) with i.p. injection of 8 mg/kg of PDRN immediately after PH. Light microscopically, liver regeneration induced by PH involved proliferation of various hepatic cells including parenchymal cells and Kupffer cells. The disintegration (1 or 2 days after PH) and remodeling (4 days after PH) of hepatic plate and increase of sinusoids (6 days after PH) were sequentially observed. These regenerative processes occurred relatively earlier in the experimental groups. By western blotting, the expression of PCNA increased in the early stage, reached its peak at 1 and 2 days after PH, and decreased thereafter. By immunohistochemistry, the intensities of PCNA staining in the experimental groups were more obvious at various days after PH. By electron microscopy using immunogold labeling for PCNA, the gold particles were largely observed in heterochromatin and nucleolus in hepatic cells. The results of immunochemistry and western blotting for PCNA showed the similar pattern in most groups. These findings suggest that PDRN treatment to hepatectomized rat could accelerate liver regeneration through rapid cell proliferation.