| Abstract |
Objectives: Staphylococcus aureus nasal carriage, an important risk factor for severe infections, is
common in patients with type 2 diabetes mellitus (T2D), possibly due to dysregulation of innate
immunity. Genetic polymorphisms in host innate immune genes, including interleukin 6 (IL6) and
vitamin D receptor (VDR) genes, have been found to be associated with susceptibility to several
diseases, including type 2 diabetes (T2D) and various infections. Additionally, vitamin D and
cathelicidin (LL-37), a downstream target of vitamin D - VDR complex, are both innate immune
markers that play a key role in immunomodulation and may contribute to host defence mechanisms
against S. aureus. The aim of this study was to investigate the role of rs1800795 IL6 single nucleotide
polymorphism (SNP) on the susceptibility to T2D, as well as to nasal carriage of S. aureus among
individuals with T2D. We also examined whether LL-37 serum levels are influenced by the four most
common VDR gene polymorphisms [FokI (rs10735810), BsmI (rs1544410), ApaI (rs7975232), and
TaqI (rs731236)] in individuals with T2D. Furthermore, we tried to determine the impact of 25-
hydroxy vitamin D [25(OH)D] and LL-37 on S. aureus nasal carriage status in T2D patients. Finally,
this study aimed to assess the contribution of S. aureus nasal colonization on subsequent
staphylococcal infections in diabetic subjects.
Patients & methods: The study group consisted of a cohort of 240 T2D patients, followed up at the
outpatient clinic of Internal Medicine and Endocrinology Department of University Hospital of
Heraklion, during 2013-2014. The control group consisted of 236 healthy age- and sex-matched
subjects. All individuals were of self-reported Cretan origin. From all T2D patients, nasal swab was
obtained to determine S. aureus nasal colonization by rotating a sterile fiber-tipped swab four times in
both anterior nares of each enrolled individual. A repeat swab was obtained, at a one-month interval,
for the estimation of persistent carriage status. Diabetic participants were asked about the use of
vitamin D supplements. Whole blood was collected from all participants enrolled in the study. One
hundred and forty-four patients with T2D and one hundred and eighty healthy controls were
genotyped for the IL6 rs1800795 SNP by Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP). FokI (rs10735810), BsmI (rs1544410), ApaI (rs7975232) and TaqI
(rs731236) VDR gene polymorphisms had previously been investigated in 173 T2D patients by PCRRFLP.
Serum levels of 25(OH)D were estimated in 118 diabetic patients and 94 controls using
chemiluminescence immunoassay (CLIA), while serum levels of LL-37 were evaluated in 118
diabetic patients using enzyme-linked immunosorbent assay (ELISA). All T2D patients were
followed up for 6 years (2014-2020) and any cases of staphylococcal infections were recorded during
this period.
Results: S. aureus nasal colonization was found to be at a percentage of 25.8% of T2D patients.
Among them, 9.6% were defined as persistent carriers. The G/C genotype and the minor allele C of
IL6 rs1800795 SNP analyzed were more common in individuals with T2D than in healthy controls
(p=0.004, OR=1.98, 95% CI 1.24-3.18 and p=0.011, OR=1.59, 95% CI 1.11-2.26, respectively). No
statistically significant differences have been found between carriers and non-carriers in terms of
either allele or genotype frequency. Analyses of subgroups, based on the pattern of carriage
(intermittent or permanent), also did not reveal significant differences. T2D patients presented
significantly lower 25(OH)D serum levels compared to controls (p<0.001). In particular, 88.6% of
T2D patients, whereas only 59.6% of controls, in our study, presented vitamin D deficiency
(<20ng/ml) or insufficiency (21-29ng/ml)). No effect of vitamin D daily administration has been
found on the rates of S. aureus nasal carriage in T2D patients (p=0.706). T2D patients presented
remarkably low serum LL-37 levels (median 0.89, range 0.05-8.62 ng/ml). The presence of the BsmI
b allele was related to higher levels of LL-37 (p=0.05), as well as BsmI bb genotype showed higher
LL-37 concentratons compared to Bb genotype (p=0.035). TaqI TT homozygotes presented higher
LL-37 levels compared to TaqI Tt heterozygotes (p=0.003), as well as the presence of the TaqI T
allele was related to a trend of increased levels of LL-37, though not statistically significant (p=0.07).
Circulating levels of LL-37 were higher in nasal carriers compared to non-carriers (p<0.001), whereas
no difference was observed in 25(OH)D levels. A strong positive correlation was observed between
25(OH)D and LL-37 among non-carriers (r=0.48, p<0.001). This relationship was independent of
vitamin D supplementation in a linear regression analysis (p=0.002). Finally, assessing the development of clinical staphylococcal infections in T2D patients during the 6-year follow-up, we
found no episodes of bacteremia, while very few episodes of skin and soft tissue infections, caused by
S. aureus, were reported.
Conclusions: Our study showed that T2D patients present low serum concentrations of vitamin D and
LL-37, suggesting defects in innate immunity, which could contribute to higher rates of S. aureus
nasal colonization. No impact of vitamin D supplementation on S. aureus nasal colonization has been
found in T2D patients. Another finding is that IL6 rs1800795 SNP, although is associated with
increased risk for T2D development, might not be associated with higher prevalence of S. aureus
nasal carriage in patients with T2D. Our findings, also, suggest that TaqI and BsmI VDR gene
polymorphisms are related to serum levels of LL-37, which could be a primary determinant for
different S. aureus carriage states in T2D. Finally, we did not prove any significant difference in the
frequency of the infections we investigated between S. aureus colonized and non-colonized patients.
A weakness of the study is the relatively small number of samples and, therefore, our results need to
be confirmed in larger studies.
|
Search
