Effects Of Propofol And Sevoflurane On Plasma Mirna Levels İn Living Kidney Donors

Following the approval of the Institutional Review Board (IRB), this prospective, comparative study will be conducted at the University Faculty of Medicine Hospital. The study will be concluded once the target number of eligible donors is reached. Written informed consent will be obtained from donors aged 18-65, classified as American Society of Anesthesiologists (ASA) physical status I-II, with a body mass index (BMI) below 35 kg/m², and scheduled for elective donor nephrectomy in the preoperative holding area before being transferred to the operating room. Informed consent forms will be obtained face-to-face by the investigators during the preoperative preparation process, prior to any clinical intervention, premedication, or randomization procedure.

Allocation of donors to the groups (Group S and Group P) will be performed in a 1:1 ratio using a computer-generated random numbers table via block randomization. To ensure strict allocation concealment, the assignments will be enclosed in sequentially numbered, opaque, sealed envelopes. The preparation of these envelopes will be handled by an independent researcher who is not involved in either the clinical phase of the study or the data analysis. Upon obtaining informed consent, the anesthesia team will open the corresponding sealed envelope in sequence and execute the assigned protocol, thereby preventing selection bias.

The Sevoflurane and Propofol methods utilized for anesthetic maintenance are part of the standard institutional routine application protocols of the Department of Anesthesiology and Reanimation. Donors will not receive any non-routine drug or dosage within the scope of this study. The randomization process is employed solely to determine which of the two standard and routine anesthetic regimens already available to the donor will be administered. Consequently, all anesthetic agents, intravenous administration sets, endotracheal tubes, and monitoring materials utilized intraoperatively are covered under the routine operational expenses of the hospital; no additional consumable materials will be introduced for research purposes.

A total of 34 donors enrolled in the study will be allocated into two groups via block randomization and the sealed envelope method to determine the maintenance regime of general anesthesia:

• Group S (Sevoflurane Group): General anesthesia maintenance will be provided using Sevoflurane at a concentration of 1.5-2% combined with a 50% Air / 50% Oxygen mixture.
• Group P (Propofol Group): General anesthesia maintenance will be provided using Total Intravenous Anesthesia (TIVA) via Propofol infusion at a dose of 4-10 mg/kg/hour combined with a 50% Air / 50% Oxygen mixture.

Sevoflurane and Propofol were selected as the comparative interventions due to their proven safety profiles and high preference rates in routine clinical practice, driven by their well-documented properties in preserving organ perfusion and ensuring rapid recovery. Both anesthetic agents are consistently and continuously stocked for routine clinical use in the hospital pharmacy. Potential adverse events, such as nausea, vomiting, shivering, or rare malignant hyperthermia associated with Sevoflurane, and injection pain, hypotension, or bradycardia associated with Propofol, will be monitored by the anesthesia team under standard monitoring and managed promptly according to institutional routine treatment protocols.

Upon arrival in the operating suite, standard monitoring-including Electrocardiography (ECG), Non-Invasive Blood Pressure (NIBP), Peripheral Oxygen Saturation (SpO2), and End-Tidal Carbon Dioxide (EtCO2)-will be established for all participating donors. To ensure pharmacological uniformity at the baseline phase between the groups, anesthesia induction will be performed using a standardized protocol in both groups as follows:

• Induction Protocol: All donors will routinely receive 0.03-0.05 mg/kg Midazolam, 2-2.5 mg/kg Propofol, 1-2 mcg/kg Fentanyl, and 0.6 mg/kg Rocuronium bromide for neuromuscular blockade, followed by endotracheal intubation.

Blood samples of 5 mL will be collected from the donors at three distinct clinical time points: prior to anesthesia induction (baseline value), at the end of surgery (post-extubation), and at the 24th postoperative hour. The blood sampling procedures will be performed during routine intravenous cannulation or through the existing intravenous line. Necessary medical precautions have been instituted to manage potential minor complications such as localized pain, hematoma, ecchymosis, or rare vasovagal syncope. The collected blood samples will be preserved by Dr. Tarik Oztuzcu at the Department of Medical Genetics, and plasma microRNA (miRNA) levels will be analyzed after the sample collection for all 34 donors is finalized.

MicroRNA Expression Methodology:

Within the scope of the project, expression levels of miR-146a, miR-155, miR-21, and the U6 gene will be analyzed in serum samples obtained by centrifuging the blood samples of the 34 donors.

• RNA Isolation: Serum samples will be used for miRNA gene expression analysis. RNA isolation will be performed in accordance with the commercial miRNeasy (Qiagen, Germany) kit protocol. Serum samples separated from blood will be lysed within QIAzol Lysis Reagent. Following the addition of chloroform, the lysate mixture will be centrifuged to separate the aqueous and organic phases. RNA will partition into the upper aqueous phase, while DNA and proteins will remain in the organic phase. The aqueous phase will be carefully transferred, and extraction steps will proceed according to the manufacturer's instructions.
• Determination of miRNA Expression: Only mature miRNAs from the isolated total RNAs will be reverse-transcribed into complementary DNA (cDNA). The miScript II Reverse Transcriptase RT kit (Qiagen, Hilden, Germany) will be used for the conversion of mature miRNAs in total RNA samples to cDNA. The reaction components and experimental steps in the cDNA synthesis phase will be carried out according to the manufacturer's protocol. Following the generation of cDNA templates from mature miRNAs, the RT-PCR array phase will be initiated. SYBR Green dye will be used along with specific primers for miR-146a, miR-155, miR-21, and U6 as an internal control to evaluate changes in the miRNA expression profile. After preparing the PCR mixture, the RT-PCR reaction will be initiated using a Corbett RT-PCR system (Qiagen, Germany). The 2^-ΔΔCt method will be utilized for relative quantification and data analysis.