Inflammation and oxidative stress transcription profiles due to in vitro supply of methionine with or without choline in unstimulated blood polymorphonuclear leukocytes from lactating Holstein cows

Abstract

Neutrophils are the most important polymorphonuclear leukocytes (PMNL), representing the front-line defense involved in pathogen clearance upon invasion. As such, they play a pivotal role in immune and inflammatory responses. Isolated PMNL from 5 mid-lactating Holstein dairy cows were used to evaluate the in vitro effect of methionine (Met) and choline (Chol) supplementation on mRNA expression of genes related to the Met cycle and innate immunity. The target genes are associated with the Met cycle, cell signaling, inflammation, antimicrobial and killing mechanisms, and pathogen recognition. Treatments were allocated in a 3 × 3 factorial arrangement, including 3 Lys-to-Met ratios (L:M, 3.6:1, 2.9:1, or 2.4:1) and 3 levels of supplemental Chol (0, 400, or 800 μg/mL). Three replicates per treatment group were incubated for 2 h at 37°C and 5% atmospheric CO 2. Both betaine-homocysteine S-methyltransferase and choline dehydrogenase were undetectable, indicating that PMNL (at least in vitro) cannot generate Met from Chol through the betaine pathway. The PMNL incubated without Chol experienced a specific state of inflammatory mediation [greater interleukin-1β (IL1B), myeloperoxidase (MPO), IL10, and IL6] and oxidative stress [greater cysteine sulfinic acid decarboxylase (CSAD), cystathionine gamma-lyase (CTH), glutathione reductase (GSR), and glutathione synthase (GSS)]. However, data from the interaction L:M × Chol indicated that this negative state could be overcome by supplementing additional Met. This was reflected in the upregulation of methionine synthase (MTR) and toll-like receptor 2 (TLR2); that is, pathogen detection ability. At the lowest level of supplemental Chol, Met downregulated GSS, GSR, IL1B, and IL6, suggesting it could reduce cellular inflammation and enhance antioxidant status. At 400 μg/mL Chol, supplemental Met upregulated PMNL recognition capacity [higher TLR4 and L-selectin (SELL)]. Overall, enhancing the supply of methyl donors to isolated unstimulated PMNL from mid-lactating dairy cows leads to a low level of PMNL activation and upregulates a cytoprotective mechanism against oxidative stress. Enhancing the supply of Met coupled with adequate Chol levels enhances the gene expression of PMNL pathogen-recognition mechanism. These data suggest that Chol supply to PMNL exposed to low levels of Met effectively downregulated the entire repertoire of innate inflammatory-responsive genes. Thus, Met availability in PMNL during an inflammatory challenge may be sufficient for mounting an appropriate biologic response.

Amino acids and the regulation of oxidative stress and immune function in dairy cattle

“…The recent work by Lopreiato et al (2019) investigated the effects of incubating bovine PMNL with Met and/or choline and observed that supplemental Met coupled with adequate choline enhanced gene expression of TLR2 and L-selectin ( SELL ), which are pathogen recognition mechanisms. In the same experiment, cells incubated without choline had greater mRNA abundance of IL1B , IL6 , IL10 , and myeloperoxidase ( MPO ), glutathione reductase ( GSR ), GSS , cystathionine gamma-lyase ( CTH ), and cysteine sulfinic acid decarboxylase ( CSAD ), indicating greater inflammation and oxidative stress; this effect, however, was ameliorated by supplementing additional Met ( Lopreiato et al, 2019 ). Thus, the increased DMI and milk production observed when feeding RPM can be partly explained by a reduction in inflammation as it directly (at the hepatic level and by dampening the immune cell overresponse) and indirectly (reducing oxidative stress) decreases circulating pro-inflammatory cytokines.…”

Role of nutraceuticals during the transition period of dairy cows: a review

“…Methionine supplementation may also have positive implications on immune cell function (Table 4) with increased phagocytosis from neutrophils, improved oxidative burst capacity, greater T-lymphocyte proliferation, and blood neutrophil-killing capacity [90,92,103]. Late pregnancy supply of methionine may have positive RPC rumen protected choline, RPM rumen protected methionine, TAG triacylglycerol, NEFA non-esterified fatty acids, PMNL polymorphonuclear leukocytes, TCA tricarboxylic acid cycle, APP negative acute phase protein, PON paraoxonase implications on calf development (Table 3) with greater calf body weight at birth and in the first weeks of life.…”

“…Methionine supplementation may also have positive implications on immune cell function (Table 4 ) with increased phagocytosis from neutrophils, improved oxidative burst capacity, greater T-lymphocyte proliferation, and blood neutrophil-killing capacity [ 90 , 92 , 103 ]. Late pregnancy supply of methionine may have positive implications on calf development (Table 3 ) with greater calf body weight at birth and in the first weeks of life.…”

Methyl donor supply to heat stress-challenged polymorphonuclear leukocytes from lactating Holstein cows enhances 1-carbon metabolism, immune response, and cytoprotective gene network abundance

“…Clearly, physiological responses in the cow to heat stress are multifaceted. Specific to immune cells, Met coupled with choline (Chol) supplementation has been associated with an improvement in PMN responses to in vitro short-term incubations and ex vivo whole-blood LPS stimulation (Vailati-Riboni et al, 2017;Lopreiato et al, 2019). The mechanisms by which supplemental Met and Chol act to modulate the immune response and oxidative stress are linked to 1-carbon metabolism, encompassing 3 pathways that represent an interconnected route through which these nutrients affect molecular events such as protein synthesis via the mechanistic target of rapamycin (mTOR), energy metabolism, and antioxidant synthesis (Coleman et al, 2020).…”