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. 2012 Oct 7;18(37):5197-204.
doi: 10.3748/wjg.v18.i37.5197.

Inhalation of Hydrogen Gas Reduces Liver Injury During Major Hepatotectomy in Swine

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Free PMC article

Inhalation of Hydrogen Gas Reduces Liver Injury During Major Hepatotectomy in Swine

Lei Xiang et al. World J Gastroenterol. .
Free PMC article

Abstract

Aim: To study the effect of H2 gas on liver injury in massive hepatectomy using the intermittent Pringle maneuver in swine.

Methods: Male Bama pigs (n = 14) treated with ketamine hydrochloride and Sumianxin II as induction drugs followed by inhalation anesthesia with 2% isoflurane, underwent 70% hepatotectomy with loss of bleeding less than 50 mL, and with hepatic pedicle occlusion for 20 min, were divided into two groups: Hydrogen-group (n = 7), the pigs with inhalation of 2% hydrogen by the tracheal intubation during major hepatotectomy; contrast-group (n = 7), underwent 70% hepatotectomy without inhalation of hydrogen. Hemodynamic changes and plasma concentrations of alanine aminotransferase (ALT), aspartate aminotransferase (AST), hyaluronic acid (HA), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and malondialdehyde (MDA) in liver tissue were measured at pre-operation, post-hepatotectomy (PH) 1 h and 3 h. The apoptosis and proliferating cell nuclear antigen (PCNA) expression in liver remnant were evaluated at PH 3 h. Then we compared the two groups by these marks to evaluate the effect of the hydrogen in the liver injury during major hepatotectomy with the Pringle Maneuver in the swine.

Results: There were no significant differences in body weight, blood loss and removal liver weight between the two groups. There was no significant difference in changes of portal vein pressure between two groups at pre-operation, PH 30 min, but in hydrogen gas treated-group it slightly decrease and lower than its in contrast-group at PH 3 h, although there were no significant difference (P = 0.655). ALT and AST in Hydrogen-group was significantly lower comparing to contrast-group (P = 0.036, P = 0.011, vs. P = 0.032, P = 0.013) at PH 1 h and 3 h, although the two groups all increased. The MDA level increased between the two group at PH 1 h and 3 h. In the hydrogen gas treated-group, the MDA level was not significantly significant at pre-operation and significantly low at PH 1 h and 3 h comparing to Contrast-group (P = 0.0005, P = 0.0004). In Hydrogen-group, the HA level was also significantly low to contrast-group (P = 0.0005, P = 0.0005) although the two groups all increased at PH 1 h and 3 h. The expression of cluster of differentiation molecule 31 molecules Hydrogen-group was low to Contrast-group. However, PCNA index (%) was not statistically significant between the two groups (P = 0.802). Microphotometric evaluation of apoptotic index (AI) in terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling-stained tissue after hepatotectomy for 3h, the AI% level in the hydrogen was significantly low to contrast-group (P = 0.012). There were no significant difference between Hydrogen-group and contrast-group at pre-operation (P = 0.653, P = 0.423), but after massive hepatotectomy, the TNF-α and IL-6 levels increase, and its in Hydrogen-group was significantly low compared with contrast-group (P = 0.022, P = 0.013, vs. P = 0.016, P = 0.012), respectively. Hydrogen-gas inhalation reduce levels of these markers and relieved morphological liver injury and apoptosis.

Conclusion: H2 gas attenuates markedly ischemia and portal hyperperfusion injury in pigs with massive hepatotectomy, possibly by the reduction of inflammation and oxidative stress, maybe a potential agent for treatment in clinic.

Keywords: Anti-oxidant; Hydrogen gas; Hyperperfusion; Malondialdehyde; Massive hepatotectomy; Oxidative stress.

Figures

Figure 1
Figure 1
Serial changes of portal vein pressure in two groups. A bar graph shows the mean ± SD of portal vein pressure (PVP) (mmHg) in the two groups. Each group is represented by the mean of 7 swines. There was no significant difference in changes of PVP between two groups at the pre-operation (pre-op), 0.5 h and 3 h.
Figure 2
Figure 2
Change of serum alanine aminotransferase level and serum aspartate aminotransferase level in two groups. Each group is represented by the mean of 7 swines. A, B: In hydrogen gas treated-group, the alanine aminotransferase (ALT) (A) and aspartate aminotransferase (AST) (B) levels were significantly lower to Contrast-group. aP < 0.05 vs ALT level in Contrast-group; cP < 0.05 vs AST level in Contrast-group. Pre-op: Pre-operation.
Figure 3
Figure 3
Changes of hepatic malondialdehyde, hyaluronic acid, tumer necrosis factor-α and interleukin-6 levels in two groups. A bar graph shows the mean ± SD of hepatic malondialdehyde (MDA) (A), hyaluronic acid (HA) (B), tumer necrosis factor (TNF)-α (C) and interleukin (IL)-6 (D) level in two groups. Each group is represented by the mean of 7 swines. A, B: In hydrogen gas treated-group, the MDA (A) and HA (B) levels were significantly lower to Contrast-group; C, D: In hydrogen gas treated-group, the TNF-α (C) and IL-6 (D) levels were significantly lower to Contrast-group. Pre-op: Pre-operation. bP < 0.01 vs MDA level in Contrast-group; dP < 0.01 vs HA level in Contrast-group; eP < 0.01 vs TNF-α level in Contrast-group; fP < 0.01 vs IL-6 level in Contrast-group.
Figure 4
Figure 4
Hematoxylin and eosin, transmission electron microscopic photographs and cluster of differentiation molecule 31 immunohistochemical staining of tissue samples taken 3 h after hepatotectomy. A: Hematoxylin and eosin (HE) staining of the Contrast-group; B: HE staining of the hydrogen gas treated-group; C, D: Transmission electron microscopic photographs of the sinusoid, arrows indicatethe sinusoidal endothelial, asterisks indicate the enlargement of the Disse's spaces; E: Cluster of differentiation molecule 31 (CD31) immunostaining of the hydrogen gas treated-group; F: CD31 immunostaining of the Contrast-group.
Figure 5
Figure 5
Proliferating cell nuclear antigen immunostaining in liver and the percentage of proliferating cell nuclear antigen stained in two groups. A: Proliferating cell nuclear antigen (PCNA) staining in liver remnant (arrows, positive cell: × 400); B: Microphotometric evaluation in PCNA stained tissue after hepatotectomy for 3 h between two groups. A bar graph shows the mean ± SD of PCNA stained level (%) in two groups. Each group is represented by the mean of 7 swines.
Figure 6
Figure 6
Terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling staining after hepatectomy and protective effect of H2 against liver apoptotic cell death in two groups. A: Revealed many terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL)-positive cells (arrows, identified morphologically by dark brown staining nuclei) in the liver remnant × 400; B: Microphotometric evaluation of apoptotic index (AI) in TUNEL-stained tissue after hepatotectomy for 3 h. A bar graph shows the mean ± SD of AI level (%) in two groups. Each group is represented by the mean of 7 swines. In hydrogen gas treated-group, the AI level was significantly low to Contrast-group (aP < 0.05 vs Contrast-group).

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