Effects of intestinal flora dysregulation on mercury concentrations in tissues and feces in methylmercury treated mice

Background  Intestinal microorganisms can directly or indirectly affect the metabolism and absorption of heavy metals, but their roles and mechanisms in the metabolism and absorption of methylmercury are still unclear.

Objective This experiment investigates the influences of intestinal flora on the absorption, distribution, and excretion of methylmercury and the possible mechanism.

Methods Twenty female BALB/c mice were randomly divided into four groups (n=5) as follows: blank group, dysbacterial group, methylmercury group, and dysbacterial+methylmercury group. Mice were given antibiotic mixture for 4 d (vancomycin 100 mg·kg^-1, neomycin sulfate 200 mg·kg^-1, metronidazole 200 mg·kg^-1, and ampicillin 200 mg·kg^-1, in body weight, thereafter, once a day, 10 mL·kg^-1) to establish an intestinal flora disorder model. The V4 region of 16S rRNA was detected by PCR to determine whether the bacterial flora disorder model was established successfully. The methylmercury group and the dysbacterial+methylmercury group were exposed to methylmercury for 3d (0.5mg·kg^-1, once a day, 10mL·kg^-1 by gavage). The feces of each group were collected 24, 48, and 72h after the first methylmercury exposure. At the end of the experiment, the mice were sacrificed, and samples of blood, liver, kidney, brain, colon, and cecum contents were collected. The total mercury levels in the liver, kidney, brain, blood, and feces were determined by atomic fluorescence spectrometry; the pH values of colon and cecum contents were tested with a pH meter; the intestinal pathological changes were observed after HE staining; the Occludin, Zo-1, and Claudin-1 (tight junction proteins) mRNA expression levels were measured by reverse transcription PCR.

Results The results of electrophoresis of the PCR amplified products of bacterial 16S rRNA universal primers under ultraviolet light showed that the mice in the blank group had good 16S rRNA gene signals, while no obvious signal was detected in the antibiotic-treated mice, indicating that the antibiotic treatment eliminated the bacteria in the mouse intestine, and the model of intestinal flora disorder was successful. After 24, 48, and 72 h of the first methylmercury exposure, the fecal mercury levels of the dysbacterial+methylmercury group(90.39±27.56), (366.75±81.82), and (641.33±354.24) ng·g^-1 were lower than the levels of the methylmercury group(259.87±90.94), (616.83±197.90), and (1322.50±377.77) ng·g^-1 (P < 0.05). The total mercury levels in the liver, kidney, brain, and blood of the dysbacterial+ methylmercury group(4 090.38±929.44), (8 539.97±1 242.64), (1 348.24±336.69), and (40 290.00±5 515.11) ng·g^-1 were higher than the levels of the methylmercury group(2 752.86±566.09), (4 502.08±105.76), (827.54±106.50), and (28 392.00±5 813.25) ng·g^-1 (P < 0.05). Compared with the mice without antibiotics treatment, the pH values of the cecum and colon contents of the antibiotic-treated mice increased (P < 0.05). HE staining results showed that the dysbacterial mice had changes such as necrosis of intestinal villi, shedding of epithelial cells, and mild edema of the lamina propria. The Occludin, Zo-1, and Claudin-1 mRNA expression levels of the antibiotic-treated mice were decreased compared with the mice without antibiotics treatment (P < 0.05).

Conclusion Intestinal flora may be an important participant in methylmercury metabolism and mercury cycle.