Differential Roles of Cystathionine Gamma-Lyase and Mercaptopyruvate Sulfurtransferase in Hapten-Induced Colitis and Contact Dermatitis in Mice.

Hydrogen sulfide (H2S) has been shown to act as both anti-inflammatory and pro-inflammatory mediators. Application of H2S donors generally protects against inflammation; however, experimental results using mice lacking endogenous H2S-producing enzymes, such as cystathionine γ-lyase (CTH) and mercaptopyruvate sulfurtransferase (MPST), are often contradictory. We herein examined two types of model hapten-induced inflammation models, colitis (an inflammatory bowel disease model of mucosal immunity) and contact dermatitis (a type IV allergic model of systemic immunity), in CTH-deficient (Cth-/-) and MPST-deficient (Mpst-/-) mice. Both mice exhibited no significant alteration from wild-type mice in trinitrobenzene sulfonic acid (Th1-type hapten)-induced colitis (a Crohn's disease model) and oxazolone (Th1/Th2 mix-type; Th2 dominant)-induced colitis (an ulcerative colitis model). However, Cth-/- (not Mpst-/-) mice displayed more exacerbated phenotypes in trinitrochlorobenzene (TNCB; Th1-type)-induced contact dermatitis, but not oxazolone, at the delayed phase (24 h post-administration) of inflammation. CTH mRNA expression was upregulated in the TNCB-treated ears of both wild-type and Mpst-/- mice. Although mRNA expression of pro-inflammatory cytokines (IL-1ß and IL-6) was upregulated in both early (2 h) and delayed phases of TNCB-triggered dermatitis in all genotypes, that of Th2 (IL-4) and Treg cytokines (IL-10) was upregulated only in Cth-/- mice, when that of Th1 cytokines (IFNγ and IL-2) was upregulated in wild-type and Mpst-/- mice at the delayed phase. These results suggest that (upregulated) CTH or H2S produced by it helps maintain Th1/Th2 balance to protect against contact dermatitis.

Increased Urinary 3-Mercaptolactate Excretion and Enhanced Passive Systemic Anaphylaxis in Mice Lacking Mercaptopyruvate Sulfurtransferase, a Model of Mercaptolactate-Cysteine Disulfiduria.

Mercaptopyruvate sulfurtransferase (Mpst) and its homolog thiosulfate sulfurtransferase (Tst = rhodanese) detoxify cyanide to thiocyanate. Mpst is attracting attention as one of the four endogenous hydrogen sulfide (H2S)/reactive sulfur species (RSS)-producing enzymes, along with cystathionine ß-synthase (Cbs), cystathionine γ-lyase (Cth), and cysteinyl-tRNA synthetase 2 (Cars2). MPST deficiency was found in 1960s among rare hereditary mercaptolactate-cysteine disulfiduria patients. Mpst-knockout (KO) mice with enhanced liver Tst expression were recently generated as its model; however, the physiological roles/significances of Mpst remain largely unknown. Here we generated three independent germ lines of Mpst-KO mice by CRISPR/Cas9 technology, all of which maintained normal hepatic Tst expression/activity. Mpst/Cth-double knockout (DKO) mice were generated via crossbreeding with our previously generated Cth-KO mice. Mpst-KO mice were born at the expected frequency and developed normally like Cth-KO mice, but displayed increased urinary 3-mercaptolactate excretion and enhanced passive systemic anaphylactic responses when compared to wild-type or Cth-KO mice. Mpst/Cth-DKO mice were also born at the expected frequency and developed normally, but excreted slightly more 3-mercaptolactate in urine compared to Mpst-KO or Cth-KO mice. Our Mpst-KO, Cth-KO, and Mpst/Cth-DKO mice, unlike semi-lethal Cbs-KO mice and lethal Cars2-KO mice, are useful tools for analyzing the unknown physiological roles of endogenous H2S/RSS production.

Preeclampsia-Like Features and Partial Lactation Failure in Mice Lacking Cystathionine γ-Lyase-An Animal Model of Cystathioninuria.

Elevated plasma homocysteine levels are considered as a risk factor for cardiovascular diseases as well as preeclampsia-a pregnancy disorder characterized by hypertension and proteinuria. We previously generated mice lacking cystathionine γ-lyase (Cth) as cystathioninuria models and found them to be with cystathioninemia/homocysteinemia. We investigated whether Cth-deficient (Cth-/-) pregnant mice display any features of preeclampsia. Cth-/- females developed normally but showed mild hypertension (~10 mmHg systolic blood pressure elevation) in late pregnancy and mild proteinuria throughout development/pregnancy. Cth-/- dams had normal numbers of pups and exhibited normal maternal behavior except slightly lower breastfeeding activity. However, half of them could not raise their pups owing to defective lactation; they could produce/store the first milk in their mammary glands but not often provide milk to their pups after the first ejection. The serum oxytocin levels and oxytocin receptor expression in the mammary glands were comparable between wild-type and Cth-/- dams, but the contraction responses of mammary gland myoepithelial cells to oxytocin were significantly lower in Cth-/- dams. The contraction responses to oxytocin were lower in uteruses isolated from Cth-/- mice. Our results suggest that elevated homocysteine or other unknown factors in preeclampsia-like Cth-/- dams interfere with oxytocin that regulates milk ejection reflex.