Upregulation of Superoxide Dismutase Activity in the Intestinal Tract Mucosa of Germ-Free Mice

Abstract

ABSTRACT. Superoxide dismutase (SOD) catalyzes the breakdown of superoxide into hydrogen peroxide and oxygen in the antioxidant defense system. We had reported that the SOD activities in the ceca of germ-free (GF) mice were significantly higher than those in conventional (CV) mice. In this study, we confirmed the location where SOD activity and protein expression increased in the ceca of GF mice. An immunohistochemical analysis and total SOD activity assay were conducted using the mucosa and other remaining tissues in the ceca. In addition to SOD activity in the ceca, 4 sites of intestinal (duodenal, jejunal, ileal and colonic) mucosae in GF mice were compared with those of CV mice. Total SOD activity in the cecal mucosa of GF mice was significantly higher than that in CV mice (P<0.01), and the intensity of CuZnSOD-positive cells in cecal mucosa was increased in all GF mice. Total and CuZnSOD activities in the duodenal, jejunal, ileal, cecal and colonic mucosae of GF mice were significantly higher than those in CV mice (P<0.05, or P<0.01). Furthermore, CuZnSOD mRNA showed similar tendencies with respect to these activities. Our results suggest for the first time that upregulation of SOD activity occurs in the entire intestinal mucosa of GF mice. Antioxidant enzymes play an important role in preventing oxidative stress by acting on reactive oxidative species (ROS) generated in vivo. While physiological concentrations of ROS in vivo are beneficial and involve cell signaling pathways and survival from invading pathogens, an unbalanced, elevated levels of ROS may contribute to the development of various diseases, such as cancer, hypertension, diabetes, atherosclerosis, inflammation, and premature aging [30]. Superoxide dismutases (SODs) [16] are the most important antioxidant enzymes in the antioxidant defense system against a ROS of superoxide (O 2•− ). At present, 3 distinct SOD isoforms have been identified in mammals. One SOD isoform has Cu and Zn in its catalytic center (CuZnSOD) and exists in the intracellular cytoplasmic compartment; the second SOD isoform has manganese (Mn) in its catalytic center (MnSOD) and exists in the mitochondria of aerobic cells; and the third isoform is an extracellular SOD [30]. SODs are the first line of defense for dismutation of excess O 2•− , which causes tissue disorders, as SODs convert O 2•− to molecular oxygen and H 2 O 2 [22]. Of the different SOD enzymes, CuZnSOD is the most abundant and widely distributed enzyme in many tissues [13].We recently reported that total SOD, CuZnSOD and MnSOD activities in the ceca of germ-free (GF) mice were significantly higher than those in the ceca of conventional (CV) mice [7]. CuZnSOD mRNA expression in the ceca of GF mice was significantly higher than that in CV mice, and CuZnSOD protein expression showed similar tendencies. Consistent with these results, the total SOD activity in conventionalized (CVz) mice decreased to the level of total SOD activity observed in the ceca of CV mice. These results suggested that the antioxidant defense ...

The Crosstalk between the Gut Microbiota and Mitochondria during Exercise

“…The absence of microbial colonization in mice was associated to a reduced levels of SCFA and serum levels of GPx and catalase after endurance swimming, which are critical antioxidants for reducing oxidative stress (Hsu et al, 2015). Different studies using mice models (Dobashi et al, 2011, 2013) have reported that the gut microbiota regulate SOD activity, yet more studies are needed to better understand the gut microbiota's role in controlling the intestinal redox balance in response to long-term intense exercise, inflammation and dysbiosis.…”

Screening for Intestinal Microflora Influencing Superoxide Dismutase Activity in Mouse Cecal Mucosa

“…Consistent with these results, the total SOD activity in conventionalized (CVz) mice decreased to the level of the total SOD activity observed in the ceca of CV mice [3]. These results suggested that the antioxidant defense system in the five sites of intestinal mucosa is influenced by intestinal microflora, which downregulates SOD activity [3, 4]. Screening for specific intestinal microflora influencing SOD activity in the mouse cecal mucosa is critical for understanding this event.…”

“…In addition, total SOD and CuZnSOD activities in the duodenal, jejunal, ileal, cecal and colonic mucosae of GF mice were significantly higher than those in the mucosa of conventional (CV) mice [4]. Consistent with these results, the total SOD activity in conventionalized (CVz) mice decreased to the level of the total SOD activity observed in the ceca of CV mice [3].…”

“…We have reported that upregulation of total SOD and CuZnSOD activities and CuZnSOD mRNA expression occurs in the mucosa of germ-free (GF) mice [3, 4]. In addition, total SOD and CuZnSOD activities in the duodenal, jejunal, ileal, cecal and colonic mucosae of GF mice were significantly higher than those in the mucosa of conventional (CV) mice [4].…”

“…The reaction product was subjected to quantitative real-time polymerase chain reaction (PCR) performed following the instructions for the 7500 Real-Time PCR System (Applied Biosystems, Foster City, CA, U.S.A.). After cDNA denaturation at 95°C for 30 sec, PCR was performed according to the following thermal cycling protocol: 95°C for 5 sec and 60°C for 34 sec in 20 µl of buffer containing SYBR Premix Ex Taq (Takara Bio Inc.) and 0.8 µ M each of forward CuZnSOD (5′-GGGTTCCACGTCCATCAGT-3′) and reverse CuZnSOD (5′-CACACGATCTTCAATGGACAC-3′) primers [3, 4]. The cDNA sequence was obtained from GenBank (accession number, NM_011434), and forward and reverse primers were designed to span different exons to avoid genomic DNA amplification.…”