Serum zinc concentration in cystic fibrosis patients

ORIGINAL ARTICLE

The Clinical Respiratory Journal

Serum zinc concentration in cystic fibrosis patients with CFTR I1234V mutation associated with pancreatic sufficiency Atqah AbdulWahab1, Ahmed Abushahin1, Mona Allangawi2, Prem Chandra3, Mohamed Osman Abdel Rahman4 and Ashraf Soliman5 1 Pediatric Pulmonology, Hamad Medical Corporation, Doha, Qatar 2 Internal Medicine, Hamad Medical Corporation, Doha, Qatar 3 Medical Research Center, Hamad Medical Corporation, Doha, Qatar 4 Laboratory Medicines and Pathology, Hamad Medical Corporation, Clinical Biochemistry Division, Doha, Qatar 5 Pediatric endocrinology, Hamad Medical Corporation, Doha, Qatar

Abstract Objective: To determine serum zinc (Zn) level among cystic fibrosis (CF) patients with homozygous CFTR I1234V mutation associated with pancreatic sufficiency (PS). Methods: A cross-sectional study was conducted including both pediatric and adult CF patients. Data on age, weight, height, body mass index (BMI), BMI Z-score, FEV1, and chronic Pseudomonas aeruginosa infection were collected. Serum Zn, albumin, and total proteins were measured and analyzed. Results: Forty-five CF patients with homozygous CFTR I1234V mutation belonging to a large Arab kindred tribe and eight CF patients with other mutations associated with pancreatic insufficiency (PI). Patient’s age ranged from 2 to 49 years with a mean age of 15.1 6 9.1 years and mean plasma Zn of 0.78 6 0.15 mcg/mL. Seven (13.2%) patients with CFTR I1234V and PS had low Zn levels (60 mmol/L) on two separate occasions in addition to the presence of two disease-causing mutations in the CFTR gene. Inclusion criteria were all of the CF patients had no respiratory infections nor exacerbation for more than 4 weeks prior to enrollment. Exclusion criteria included all CF patients who were vegetarian, those with significant liver disease as defined by clinical findings of portal hypertension or cirrhosis or AST, ALT, or GGT > 2 the upper limit of normal, gall bladder disease, diabetes mellitus requiring insulin therapy, renal failure, malignancies, or collagen vascular disease or taking medications known to have an effect zinc status such as oral contraceptives or use of oral immunosuppressive drugs (steroids and nonsteroid antiinflammatory drugs). This study was reviewed and approved by the local ethics committee at Hamad Medical Corporation (Ref#12226/12). All CF patients were assessed based on the medical history including frequent pulmonary exacerbation, and whether their lower lung secretions harbor chronic P. aeruginosa colonization.

Anthropometric measurements Anthropometric measurements were performed using digital electronic platform scale and standing height measurement without shoes using a stadiometer. Body mass index (BMI) was calculated by dividing weight in kg by height squared in meters {weight (Kg)/(height (m)) 2}. The standard deviation [Z] score for BMI was calculated and adjusted for age and gender. Spirometry was obtained by standard methods in patients old enough to perform adequately (24). The highest of three technically appropriate measurements was recorded. Forced vital capacity (FVC; in liters) and (% predicted), forced expiratory volume in 1 s (FEV1; in liters) and (% predicted), and FEV1/FVC were measured using a flow-sensing spirometer (Sensor Medicus Model V6200, Germany).

The Clinical Respiratory Journal (2015) • ISSN 1752-6981 C 2015 John Wiley & Sons Ltd V

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Biochemical analysis Biochemical analysisfor serum Zn: 5 mL of whole blood in trace metal free tube was collected from participants (non fasting blood sample). After clotting and centrifugation, serum was stored at 2608C until analysis. Zn levels were determined by Flame Atomic Absorption spectrophotometer using Thermo electron corporation analyzer. The tests performed by an experience senior laboratory technician. Results were compared to healthy controls from the same tribe. The normal reference range for Zn is 0–10 years: 0.60–1.20 mcg/mL and for > or =11 years: 0.66–1.10 mcg/mL (25, 26). Albumin and total proteins were measured using Bromcresol Green method (BCG Method).

Statistical analysis Categorical data were expressed as a frequency along with percentage and continuous data values presented in mean 6 standard deviation and median and range. Descriptive statistics were used to summarize all demographic anthropometric, serum Zn, and spirometric measures and other characteristics of the participants. Unpaired t and Mann–Whitney U-tests were applied to compare mean of quantitative variables between the two independent groups (normal Zn levels and deficient Zn levels). Associations between two or more qualitative variables were assessed using chi-squared and Yates corrected chi-squared tests. Pearson’s correlation coefficient was used to assess the strength of linear relationship between two or more quantitative variables. A two-sided P value 0.05) (Fig. 1). The mean height SDS was 20.76 6 0.97 and BMI Z-score was 0.59 6 1.4. Thirty-nine patients were able to perform spirometry adequately, that include all the seven CF patients with

Figure 1. Relationship between age and zinc.


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Zn-deficient and 32 CF patients with normal Zn level. The mean FEV1 (% predicted) was 79.1 6 22.2 and the mean FVC (%predicted) was 83.4 6 20.6%. All CF deficient Zn were not taking multivitamins. Patients mean age in the Zn deficient group was significantly higher, 23.0 6 12.2 years compared to that observed in the normal Zn levels group, 13.9 6 8.1 years (P 5 0.013) in Table 2. The mean albumin was observed to be higher among zinc sufficient group compared to Zn deficient group (43.8 6 3.5 vs 40.4 6 3.6; P 5 0.027). The mean FEV1 (% predicted) in the Zn deficient group was found to be lower, 66.5 6 29.6% compared to patients with normal Zn levels, 81.4 6 20.4% (P 5 0.133). Persistent P. aeruginosa colonization was more prevalent in Zn deficient group. BMI Z-score, FEV1%, FVC%, and FEV1/FVC were positively associated with Zn levels. Forty-five healthy subjects belonging to the same Arab tribe were selected in order to assess their Zn levels and their mean plasma Zn of 0.84 6 0.11 mcg/mL (range 0.65–1.1 mcg/mL) with mean age 20.4 6 10.1 years (range 6–40 years).

Discussion In this study, we evaluated the zinc status in a selected group of CF patients associated with PS. Hypozincemia was diagnosed in seven CF patients with homozygous CFTR I1234V mutation, although, they did not have any specific symptoms of Zn deficiency. However, they had more frequent pulmonary exacerbations and more frequently harbor pseudomonas colonization. Research studies in humans and in cell models point to the useful effect of Zn in reducing inflammatory

cytokines at the mRNA level and involves inhibition of NF-kB activation (27). P. aeroginosa in subjects with CF and in CF animal models can lead to activation of the NF-kB pathway and overproduction of adhesion molecules, inflammatory cytokines (IL-1b, TNF-a, IL6, and IL-8), and oxidative stress molecules, which in turn continue to provide stimulus for further NF-kB activation and the relentless vicious cycle (28–30). A recent study provided evidence that 30 mg/day supplemental Zn was associated with reduction in the number of days of oral antibiotics used to treat respiratory tract infections and marginal reduction in the level of plasma inflammatory cytokines in children with CF (31). A number of investigators have reported conflicting relationships between Zn status, malabsorption, impaired growth, and pulmonary function in CF patients (16–19, 32, 33). Few studies found no correlation between Zn levels and nutritional or pulmonary status (17, 18). Others reported abnormal Zn status in certain patients with CF especially those with moderate to severe growth retardation and severe pulmonary disease as compared with those without growth failure and with moderate pulmonary disease (33). In this study, there were association between Zn levels and BMI Z score and FEV1, however, it was not statistically significant (P > 0.05). Zn deficient CF patients have lower BMI Z score, FEV1 and harbor more P. aeruginosa colonization compared to those with normal Zn levels. In a recent retrospective study reported the occurrence of Zn deficiency in CF adults, highlighting the fact that CF subjects with low Zn status exhibit worse pulmonary

Table 2. Association of demographic and clinical characteristics between low and normal zinc levels Variables

Zn < 0.6 mcg/mL N57

Zn 0.6 mcg/mL N 5 46

P value

Age (years) Male Height SDS BMI Z score Spirometry (yes) FEV1% Albumin g/L Total protein g/L Homozygous CFTR I1234V mutation Pancreatic sufficiency (yes)* Enzyme replacement (yes)* Multivitamin Intake (yes)* Persistent P. aeruginosa (yes)*

23.0 6 12.2 [19.8; 12–49] 5 (71.4) 20.50 6 1.1 [20.67; 21.7–1.0] 0.22 6 1.8 [20.06; 21.8–3.8] 7 (100%) 66.5 6 29.6 [73.5; 30–98] 40.4 6 3.6 [39; 35–46] 74.0 6 2.8 [73; 70–78] 7 (100) 7 (100) 0 (0) 0 (0) 6 (85.7)

13.9 6 8.1 [13.1; 2–33] 26 (56.5) 20.80 6 0.96 [20.85; 22.7–1.2] 0.65 6 1.4 [0.31; 21.19–4.62] 32 (69.6%) 81.4 6 20.4 [80; 20–113] 43.8 6 3.5 [44; 33–50] 73.3 6 5.8 [72.5; 62–88] 38 (82.6) 38 (82.6) 8 (17.39) 16 (34.8) 13 (28.3)

0.013 0.686 0.451 0.462 0.089 0.133 0.027 0.763 0.251 0.251 0.748 0.154 0.011

Quantitative values were expressed in mean 6 standard deviation and [median; range]. Qualitative values were expressed in frequency and percentage; n (%). *Yates corrected chi-squared P value. SDS, standard deviation score.

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function and a higher prevalence of impaired bone and glycemic status but they did not report whether these patients were colonized with P. aeruginosa colonization or other organisms (34). It is to be noted that that the primary analysis was not to determine the predictive values of different confounders or related factors associated with serum Zn levels and also due to the limited number of patients in the Zn deficient group, exploration for any multivariate regression analysis could not be possible and performed. Because Zn plays an important role in immunity, the decreased serum Zn in patients with decreased pulmonary function could enhance infection, leading to a vicious circle or could be a result of decreased immunity in Zn deficient CF patients that may lead to frequent pulmonary exacerbation specialty among those with chronic P. aeruginosa leading to decreased pulmonary function. Eight PI CF patients in this study had normal Zn levels and this may be related to multivitamin intake containing Zn and on PERT that improves Zn absorption. The group of Hambidge (35) showed a decreased fractional absorption of Zn in PI CF though pancreatic enzyme replacement has been shown to improve zinc absorption in children (35). The limitation of this study is the low numbers of CF patients. However, an important finding is that seven zinc deficient CF patients were all PS and all the eight PI CF patients had normal Zn levels.


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Conclusion Zn deficiency can occur in patients with CF and PS and appeared not to be associated with PI. Further investigations are needed to understand the role of Zn in mediating pulmonary function in CF subjects and to explore the association between zinc status and clinical outcomes of CF patients such as cystic fibrosis related diabetes, bone, kidney, and liver diseases.

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