Chromium picolinate (CrPic) is a mineral chelate used in human and ..... Lien TF, Yang KH, Link KJ (2005) Effects of chromium propionate supplementation on ...
Biol Trace Elem Res DOI 10.1007/s12011-013-9731-7
Humoral and Cellular Immunity in Chromium Picolinate-Supplemented Lambs B. S. L. Dallago & C. M. McManus & D. F. Caldeira & A. Campeche & R. T. Burtet & T. P. Paim & E. F. Gomes & R. P. Branquinho & S. V. Braz & H. Louvandini
Received: 9 April 2013 / Accepted: 6 June 2013 # Springer Science+Business Media New York 2013
Abstract The effects of oral supplementation of chromium picolinate (CrPic) on humoral and cellular immunity in sheep were investigated. Twenty-four male lambs divided into four treatments and received different dosages of CrPic: placebo (0), 0.250, 0.375, and 0.500 mg of chromium/animal/day during 84 days. The base ration was Panicum maximum cv Massai hay and concentrate. Blood samples were collected fortnightly for total and differential leukocyte counts. On days 28 and 56, the lambs were challenged with chicken ovalbumin I.M. Serum samples were collected on days 46 and 74 and subjected to an indirect enzyme-linked immunosorbent assay to measure IgG anti-ovalbumin. The cellmediated immune response was determined by a delay-type hypersensitivity test using phytohemagglutinin. CrPic did not significantly affect humoral immunity in lambs but there was a negative effect on cellular immunity (P0.05)
c
Except on day 56
then submitted to a chloridric digestion according to Silva and Queiroz [16]. Data were analyzed using a polynomial regression (linear and quadratic) with the Statistical Analysis System (SAS® v.9.3 Cary, NC, USA) using a completely randomized design with total Cr levels as the independent source of variation. For leukocyte count and differential leukocyte count, a repeated-measures analysis of variance was carried out. Means were then compared using multiple comparison adjustment to Tukey’s test (p0.05)
c
Measured in absorbance of 405 nm
Dallago et al. Fig. 1 Relationship between skin thickness after PHA injection (DTH) and time after treatment in control (y=0.0086x+5.96 (R2 =0.78)) and CrPic-supplemented lambs (0.250 mg of CrPic/day, y=0.0063x+5.40 (R2 =0.82); 0.375 mg of CrPic/day, y=0.0055x+5.35 (R2 =0.62); and 0.500 mg of CrPic/day, y=0.0041x+5.62 (R2 =0.7))
differences between treatments in the analysis of variance: as dietary Cr content increases, lower response to the DTH test is seen (P=0.023).
Discussion With the exception of a few studies [10, 17–20], the study of Cr supplementation in livestock industry was been relatively overlooked in the last decade, resulting in a lack of new information on this topic. This may be due to the intrinsic difficulty of working with this micromineral or may be related to the debatable results observed to date. Despite its controversial role in animal nutrition and the lack of scientific support for its use, Cr is still added by the industry as an infallible constituent in salt formulas to “improve performance and the immune system” and as a “nutritional anti-stress factor” as seen with the advertising of these products. Stressful situations have been linked to a beneficial role for Cr [15, 21, 22]. This pattern maybe true for some Fig. 2 Relationship between levels of CrPic consumed and slopes from straight lines generated after skin thickness sensitization by PHA. There is a significant difference between treatments (P=0.023) as seen by the straight line (y=−0.0065x+0.008; R2 =0.28)
parameters, such as performance [1, 7], but there is no consensus of its effect in improving immune function. In the present study, adding chromium to animals under normal conditions (without stress) was shown to be ineffective and even harmful in terms of immunity, leading to an additional superfluous cost within the production system. The differences between treatments for differential leukocyte counts on day 56 do not represent an important finding because there was no specific profile throughout time. In addition, the few alterations in total and differential leukocyte count observed indicate that CrPic supplementation do not present a direct effect on white cell production. However, these results do not exclude the possibility of impairment of white cell function. The DTH results showed impairment of some white cell functions, especially on T-lymphocyte mobilization capacity: PHA specifically links to glycoproteins on the T-lymphocyte surface resulting in their activation [23]. Once activated, the T cells release many signals (cytokines) to begin the cellular immunity effector phase, especially macrophage activation [23]. This process takes some time to
Immunity in Chromium Picolinate-Supplemented Lambs
occur—so the name “delay-type hypersensitivity”—and explains the time effect observed for this parameter. As PHA is a specific T-lymphocyte mitogen, and animals supplemented with CrPic showed a lower reaction when compared with control animals (P=0.0076), it seems that CrPic impairs Tlymphocyte action. A study conducted by Burton et al. [21] reinforces (and maybe can help to explain) the DTH results found here: the mononuclear cells from dairy cattle previously supplemented with 0.5 mg of Cr/day produced low concentrations of IL-2, INF-γ, and TNF-α after a stimulation with other mitogen, concanavalin A. TNF-α and INF-γ are the major substances responsible for the leakage of plasmatic macromolecules that cause skin toughening and thickness increases in the cutaneous delay-type hypersensitivity. Thus, they are directly involved in the cellular immunity response played by TH1 cells [23]. Both experiments reveal a probable immunosuppressive effect of Cr acting on T cell differentiation or activation, especially on TH1 cells. Regarding humoral immunity, although many studies using other animal species as models supported Cr capacity to influence antibody production, only few of them tried to explain how it acts. This has led to inconsistency between studies: some indicate that using Cr is beneficial [1, 12, 24] while others show no improvement in any parameter measured [25] or even impairment in some aspects [7]. It looks like other factors (maybe the kind of antigen used, the type and intensity of stressful situation to which the animal is submitted, or even the chemical form of supplemental Cr) is involved in modulating the humoral immunity response. In the present study, CrPic did not modify the action of B cells as no differences between treatments were observed. On the other hand, Burton et al. [24] observed increased antibody titers to ovalbumin in dairy cattle (on peripartum or early lactation) receiving 0.5 mg of Cr chelate/day but, in the same animals and under the same conditions, no differences in antihuman erythrocyte antibody production was seen. On the other hand, Moonsie-Shageer and Mowat [1] found a high antibody titre against human erythrocytes 14 days after the challenge in heifers submitted to a journey (about 26 h) and treated with 0.2, 0.5, and 1 mg of Cr/kg in the diet. Nevertheless, in weanling pigs supplemented with 0.2 mg of chromium propionate, higher titres for sheep red blood cells [26] were seen. A challenge with Escherichia coli lipopolysaccharide (0.1 mg/kg BW) was used in this last experiment as a stress-inducing agent. In addition, one trial using cattle submitted to a short trip (less than 100 km) showed no effects on IgM and IgG production against Pasteurella hemolytica after Cr supplementation in the form of CrCl3 or yeast, but when supplementation was by chromium nicotinate (CrNic), IgG production increased [12]. These previous studies confirm the controversial results obtained using supplementary Cr to date.
In spite of these results, according to Mallard et al. [22], the differences between results of Cr studies for specific antibody production have an antigen-dependent behavior in which some features such as differences between the kinds of antigen, dose given, adjuvant used, or the previous exposure to antigen interfere with antibody production. All these variables influence the evaluation of humoral immunity, but with a natural challenge in the field, the same components are relevant and are thought to be involved in individual resistance to infection. So, although comparison of results between studies is still difficult, the results are valid and must be evaluated carefully.
Conclusion Oral chromium supplementation using CrPic did not affect humoral immune response in lambs, but it impaired the T cell mobilization capacity. More studies using Cr supplements are necessary to ensure its safe use, especially in terms of for how long it should be used and whether its use is really beneficial. For the present, dietetic Cr supplementation should be avoided in nonstressed animals. Acknowledgments The authors would like to thank CNPq, INCTPecuária (CNPq/FAPEMIG), and CAPES/PROCAD-“Novas Fronteiras 2007” for the scholarships and to FINATEC and FAPDF for financial support. Special thanks to the teachers Márcio Martins Pimentel and Andrea Queiroz Maranhão for their technical support of this study.
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