الفهرس 
PERIPHERAL BLOOD MONONUCLEAR CELLSOnly 14 pages are availabe for public view
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Abstract
I
nfection in chronic kidney disease patients (CKD) is a challenging medical problem; it is the second most common cause of morbidity and mortality among CKD patients and accounts, together with cardiovascular diseases, for around 60% of mortality among end stage kidney disease (ESKD) patients.
Uremia, among other causes, was being investigated as a direct cause of the immunocompromised status among CKD patients throughout the stages of their kidney disease.
Klotho gene, the anti-aging gene, was discovered in 1997. It encodes two subtypes of klotho proteins: transmembrane protein (α, β and γ klotho) and soluble klotho. Since the discovery of Klotho, it has been a target for many researchers on animals and humans, to reveal its functions on the performance and vitality of human cells.
Alpha Klotho (α-KL) is an obligate coreceptor for fibroblast growth factor 23 (FGF23) on renal tubular cells to properly perform its well-known role in mineral metabolism. And as mineral and bone disorders are well-established among CKD patients, Klotho/ FGF23 axis is well-known to have a pivotal role among CKD patients.
Recently Klotho/ FGF23 axis malfunction has been investigated as a potential cause of the immunocompromised status among ESKD patients; but the exact function of Klotho/ FGF23 axis on the immune cells is still unknown. This hypothesis is highly supported by the discovered evidence of the anti-inflammatory functions of Klotho protein.
In the present study, we aimed to measure the expression of -klotho protein on peripheral blood lymphocytes (PBL) among hemodialysis patients as a possible contribution to their immunocompromised status, in comparison to a group of normal healthy individuals.
This case-control study included 20 ESKD patients on regular hemodialysis for at least 3 months. Their ages ranged from 24 to 69 years. All patients were recruited from Ain Shams University Dialysis Center from December 2021 till April 2022.
Patients with primary immunodeficiencies, those on systemic immunosuppressive drugs, those with ongoing infections or who had recently recovered from infections, and those with malignancies on active treatment were excluded. Twenty healthy subjects of comparable age and gender were also included as the control group.
The study was examined and approved by the Faculty of Medicine Ain Shams University Research Ethical Committee (FMASU REC), Cairo, Egypt (number: FMASU MD21/2021). A written informed consent was signed by all participants prior to their participation in the study.
Full detailed medical history and physical examination were performed for all participants.
Blood samples were collected and the following laboratory investigations according to the study protocol were performed:
• Routine laboratory investigations: complete blood picture with a differential count, urea, creatinine, AST, ALT, serum albumin and C-reactive protein.
• Peripheral blood mononuclear cells were isolated from fresh blood samples and stored at -80°C until analysis.
• The expression of α-Klotho protein was measured on three populations of lymphocytes: B-cell (CD19+), T-cell (CD3+), and NK cells (CD56+) using multi-colour flow cytometry (FCM) analysis. The results were presented as percentage of gated cells.
In our study, both groups were comparable regarding age, gender, and smoking status.
The underlying cause of CKD was identified in most cases; however, in 30% of cases, the cause was unknown. In the hemodialysis group, 25% of cases had hypertensive nephrosclerosis. Regarding the associated comorbidities, most hemodialysis patients were hypertensive, whereas patients with diabetes mellitus, cardiovascular diseases, and cerebrovascular diseases constituted the minor group among patients.
Significantly lower numbers of gated lymphocytes were detected by flow cytometry among hemodialysis patients as compared to healthy controls. We also noticed a significant reduction in the percentages of natural killer cells among hemodialysis patients.
We demonstrated significant reductions in percentages of α-KL protein expression on B lymphocytes (CD19), T lymphocytes (CD3), and natural killer cells (CD56) among hemodialysis patients compared to the control group.
At a cut-off value of 0.65, 39.0, and 8.2; the percentages of CD19/ α-Klotho+, CD3/ α-Klotho+, and CD56/ α-Klotho+ cells, respectively, had a sensitivity ranging from 80% to 85% and a specificity ranging from 90 to 95% to discriminate hemodialysis patients from healthy controls.
The present study suggests that decreased α-KL expression on PBLs may contribute to the immunocompromised status among hemodialysis patients, highlighting the importance of understanding the exact function of α-KL protein on immune cells. This may offer a future diagnostic and therapeutic tool to improve the immune response among hemodialysis patients.
CONCLUSION
from the present study we conclude the following:
• Lymphopenia is observed among hemodialysis patients as compared to healthy controls.
• The expression of α-klotho protein on B lymphocytes, T lymphocytes, and natural killer cells is significantly reduced among hemodialysis patients as compared to healthy controls.
• We assume that the decreased α-klotho protein expression on peripheral blood lymphocytes may cause an α-klotho/FGF23 axis defect and malfunctioning lymphocytes.
• Decreased α-klotho expression on immune cells may contribute to the immunocompromised status among hemodialysis patients.
RECOMMENDATIONS
from the present study we recommend the following:
• Further large-scale studies are required to explain the molecular mechanism by which α-klotho protein affects the function of lymphocytes, and how it may act as a prognostic and a therapeutic tool among CKD and hemodialysis patients.
• Further longitudinal studies are required to study α-klotho protein expression on immune cells in relation to the duration of hemodialysis.
• Further prospective studies are required to study α-klotho protein expression on immune cells in different CKD stages.