Daily preventive zinc supplementation decreases lymphocyte and eosinophil concentrations in rural Laotian children from communities with a high prevalence of zinc deficiency: results of a randomized controlled trial

Authors: KEWCHAROENWONG, CHIDCHAMAI - Khon Kaen University; SCHUSTER, GERTRUD - University Of California, Davis; WESSELLS, RYAN - University Of California, Davis; HINNOUHO, GUY-MARINO - University Of California, Davis; BARFFOUR, MAXWELL - University Of California, Davis; KOUNNAVONG, SENGCHANH - Tropical Medicine Research Institute; BROWN, KENNETH - University Of California, Davis; HESS, SONJA - University Of California, Davis; SAMER, WARAPORN - Khon Kaen University; TUSSAKHON, INTHIRA - Khon Kaen University; LERTMEMONGKOLCHAI, GANJANA - Khon Kaen University; PEERSON, JANET - University Of California, Davis; Stephensen, Charles

Submitted to: Journal of Nutrition
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/28/2020
Publication Date: 5/2/2020
Citation: Kewcharoenwong, C., Schuster, G.U., Wessells, R.K., Hinnouho, G., Barffour, M.A., Kounnavong, S., Brown, K.H., Hess, S.Y., Samer, W., Tussakhon, I., Lertmemongkolchai, G., Peerson, J.M., Stephensen, C.B. 2020. Daily preventive zinc supplementation decreases lymphocyte and eosinophil concentrations in rural laotian children from communities with a high prevalence of zinc deficiency: results of a randomized controlled trial. Journal of Nutrition. 150(8):2204-2213. https://doi.org/10.1093/jn/nxaa037.
DOI: https://doi.org/10.1093/jn/nxaa037

Interpretive Summary: Zinc deficiency, which is common in young children in many areas of the world, can impair immune function and increase the risk or severity of childhood infectious diseases. The objective of this study was to investigate the effect of three different zinc supplementation regimens, and a placebo control, on the immune system of young children in Laos, where zinc deficiency is common. In total, 512 children aged 6 to 23 months were enrolled into this study and randomly assigned to receive one of four treatments for nine months: (1) daily preventive zinc tablets (PZ; 7 mg/d); (2) daily multiple micronutrient powder to be added to meals (MNP; containing 10 mg/d zinc, 6 mg/d iron and other 13 nutrients); (3) therapeutic dispersible zinc tablets (TZ; 20 mg/d) for 10 d during episodes of diarrhea only (as zinc can decrease the duration of diarrhea); and (4) daily placebo powder (PL). To assess immune function, study investigators measured the production of cytokine mediators of inflammation from whole blood cultures, and the concentrations of specific white blood cells in peripheral blood, including neutrophils, eosinophils, basophils, monocytes, lymphocytes and a subset of lymphocytes (T lymphocytes). Study investigators hypothesized that zinc supplementation would increase cytokine production and T lymphocyte concentrations. The PZ and MNP treatments improved zinc status, as indicated by an increase in plasma zinc, compared with the PL group over the study period. However, even with improvements in zinc status in two groups, the production of cytokine mediators and of most specific types of white blood cells. Including T lymphocytes, were not affected by any of the three zinc treatments. The two exceptions were lymphocytes and eosinophils. The PZ treatment caused a significant decrease in lymphocytes relative to the PL group, which was an unexpected result that could suggest that zinc was improving an underlying infection that caused an increase in lymphocytes. The PZ treatment also caused a significant decrease in eosinophils relative to the PL group, though only in children with low plasma zinc levels at the beginning of the study, indicating that they were the most deficient. This decrease in eosinophils may also have been due to the zinc improving an underlying condition that had caused an increase in blood eosinophil levels, perhaps an undiagnosed infection. In summary, zinc supplementation did not improve immune function as hypothesized but did affect lymphocyte and eosinophil concentrations in an unexpected manner. Further work is needed to confirm that these effects of zinc on lymphocytes and eosinophils are reproducible, and to determine why this occurs.

Technical Abstract: Objectives: To investigate the effect of optimal zinc supplementation on innate and adaptive immunity in young Laotian children. Study design: This study on immune function was conducted with a subset of participants in a randomized controlled trial examining the effects of different zinc supplementation strategies for improving growth and hematologic and micronutrient status in rural Laotian children. In total, 512 children aged 6 to 23 months were enrolled for this study and randomly assigned to receive either daily preventive zinc tablets (PZ; 7 mg/d), daily multiple micronutrient powder (MNP; containing 10 mg/d zinc, 6 mg/d iron and other 13 nutrients), therapeutic dispersible zinc tablets (TZ; 20 mg/d) for 10 d for episodes of diarrhea, or daily placebo powder (PL) and followed for 9 months. Cytokine production from whole blood cultures, the concentrations of T-cell populations and a complete blood count (CBC) with differential count for leukocytes were measured at baseline and endline of the rural Laotian children. Means of endline values were compared with analysis of covariance, controlling for the baseline value of the outcome, child age and sex, district, month of enrollment, and zinc status. Results: The PZ and MNP treatments caused a significant increase in plasma zinc compared with the PL group over the study period. T-cell cytokines (IL-2, IFN-', IL-13, IL-17), LPS-stimulated cytokines (IL-1ß IL-6, TNF-a, and IL-10) and T-cell concentrations (total and memory CD4 and CD8 T cells, and CD4 Treg cells) at endline adjusted for baseline concentration showed no difference among these treatment groups, nor was there an interaction with baseline zinc status. However, CBC data revealed that endline lymphocyte concentrations were significantly lower in the TZ than PL group (p=0.032). Interactions of treatment with baseline zinc status were seen for eosinophils (pixn=0.0036), basophils (pixn=0.023) and monocytes (p = 0.086) but a significant difference between groups was seen only for eosinophils, where concentrations were significantly lower in the TZ than PL group in children with baseline plasma zinc below the overall median (p=0.012). Conclusions: Zinc supplementation regimens had no effect on innate and adaptive cytokine productions, and T-cell concentrations; nevertheless, an apparent effect of zinc was found on total lymphocytes and eosinophils. Thus, examining these cell subsets may prove to be useful as immunologic indicators in response to zinc supplementation.