#781 Cerebral vessels and early diabetic kidney disease in type 2 diabetes mellitus patients share a common mitochondrial DNA and inflammatory profile
Authors: Petrica Ligia , Gadalean Florica , Vlad Daliborca , Popescu Roxana , Bob Flaviu , Milas Livia Oana , Glavan Mihaela , Jianu Catalin , Ursoniu Sorin , Mogos Maria , Ienciu Silvia , Vlad Adrian 
Published: May 2024

Abstract

Background and Aims
Cerebrovascular disease accounts for major motor and cognitive disabilities in patients with type 2 diabetes mellitus (DM). Among the multiple pathogenic pathways which involve both the kidney and the brain vasculature, inflammation and mitochondrial dysfunction are of major importance. The aim of the study was to evaluate the association of mitochondrial DNA (mtDNA) modifications in blood and urine and of inflammatory markers with cerebral vessels structural and functional modifications in patients with type 2 DM and early diabetic kidney disease (DKD).

Method
A cohort of 150 patients (52 patients with normoalbuminuria, 48 patients with microalbuminuria, and 50 patients with macroalbuminuria) and 30 age- and gender-matched healthy controls were enrolled in this case series study. All patients were assessed concerning urinary albumin/creatinine ratio (UACR), biomarkers of podocyte damage (synaptopodin, podocalyxin) and of proximal tubule dysfunction (kidney injury molecule-1-KIM-1, N-acetyl-β-(D)-glucosaminidase-NAG), serum and urinary pro-inflammatory interleukins IL-17A, IL-18, tumor necrosis factor alpha (TNFα), intercellular adhesion molecule-1 (ICAM-1), anti-inflammatory IL-10, and eGFR (creatinine-cystatin C).
MtDNA-CN and nuclear DNA (nDNA) were quantified in peripheral blood and urine by qRT-PCR (ABI 7900-HT–Applied BioSystem). TaqMan assays were utilized for the assessment of cytochrome b (CYTB) gene, subunit 2 of NADH dehydrogenase (ND2), and of beta 2 microglobulin nuclear gene (B2M). mtDNA-CN was defined as the ratio of the number of mtDNA/nDNA copies, through analysis of the CYTB/B2M and ND2/B2M ratio.
The cerebrovascular hemodynamic indices were evaluated by an ultrasound equipment with high resolution (MYLAB 8-ESAOTE Italy) equipped with a Colour Ultrasound System. Subclinical atherosclerosis and arteriosclerosis were assessed by intima-media thickness (IMT) in the common carotid arteries (CCAs), Gosling's pulsatility index (PI) and Pourcellot's resistivity index (RI) in the internal carotid arteries (ICAs) and middle cerebral arteries (MCAs), bilaterally. Cerebrovascular reactivity (CVR), which represents the vasodilatory reactivity of cerebral vessels in response to a vasodilatory stimulus, was assessed by the breath-holding test where hypercapnia was the vasodilatory stimulus and the breath-holding index (BHI) was calculated.

Results
Multivariable linear analysis revealed direct correlations of serum TNFα with UACR, KIM-1, NAG, and urinary mtDNA, and a negative correlation with serum mtDNA (R2 = 0.893; p &lt; 0.0001); urinary TNFα correlated directly with UACR, KIM-1, NAG, and urinary mtDNA (R2 = 0.693; p &lt; 0.0001). Serum ICAM-1 had a direct correlation with UACR, KIM-1, NAG, synaptopodin, and podocalyxin, and a negative one with serum mtDNA (R2 = 0.855; p &lt; 0.0001). Urinary ICAM-1 correlated directly with UACR, KIM-1, NAG, and urinary mtDNA, and indirectly with eGFR (R2 = 0.783; p &lt; 0.0001).
CCA-IMT had a direct correlation with serum IL-17A and indirect correlations with serum mtDNA and serum IL-10 (R2 = 0.706; p &lt; 0.0001); ICA-PIs and MCA-PIs correlated directly with serum IL-17A and negatively with serum IL-10, respectively (R2 = 0.567; R2 = 0.642; p &lt; 0.0001). ICA-RIs showed a direct correlation with UACR, serum and urinary IL-17A, serum and urinary IL-18, NAG, and urinary TNFα, and a negative correlation with serum mtDNA and with serum and urinary IL-10 (R2 = 0.930; p &lt; 0.0001). MCA-RIs correlated directly with UACR, serum IL-17A, serum IL-18, NAG, urinary mtDNA, and urinary TNFα, and negatively with serum mtDNA and serum IL-10 (R2 = 0.875; p &lt; 0.0001). BHI correlated directly with serum IL-10 and serum mtDNA, and negatively with serum IL-17A, serum ICAM-1, and NAG (R2 = 0.823; p &lt; 0.0001).

Conclusion
In neurologically asymptomatic type 2 DM patients there is an association of mtDNA changes in blood and urine with cerebral vessels atherosclerotic and arteriosclerotic remodeling, as well as with decreased CVR. MtDNA modifications may exert their effect in parallel with markers of inflammation at both kidney and cerebrovascular level from the early stages of DKD.