İnsan trofoblast hücre hattinda sialik asit bağlayici reseptörlerin (siglecs) ekspresyonu

Year 2024, , 195 - 203, 01.04.2024

Nazlı Çil , İ.veysel Fenkci , Gülçin Abban Mete , Doğukan Mutlu , Cihan Kabukçu , Ümit Çabuş

https://doi.org/10.31362/patd.1346146

Abstract

Amaç: Siglecler olarak bilinen hücre yüzeyi proteinleri, sialik asit için özel bir afinite sergilemektedir. Öncelikle bağışıklık hücrelerinin yüzeyinde bulunan Siglec'ler, I-tipi lektinler adı verilen lektin alt kümesine aittir. Siglecler maternal-fetal immün toleransta önemli rollere sahiptir. Bu çalışmada İnsan Villöz Trofoblast (HVT) hücre hatlarında Siglec tiplerinin belirlenmesi amaçlanmıştır.
Gereç ve yöntem: HVT hücre hattından total RNA ekstrakte edildi, cDNA sentezlendi ve Siglec -3, -5, -6, -7, -8, -9, -10, -11 ve -16 gen ekspresyonu Gerçek Zamanlı Kantitatif Ters Transkripsiyon Polimeraz Zincir Reaksiyonu (qRT-PCR) ile analiz edildi. Ek olarak, Siglec seviyelerini belirlemek için immünohistokimyasal boyamalar yapılmıştır. Siglec-6 ve Siglec-9' a karşı immünoreaktivite kahverengi rengin yoğunluğuna göre ayrı ayrı değerlendirilmiştir.
Bulgular: HVT hücre hattından ekstrakte edilen RNA kullanılarak Siglec genlerinin qRT-PCR ile ekspresyon seviyeleri ile erime eğrisi analizleri yapıldı. Siglec -3, -5, -6, -7, -9, -10, -11 ve -16 genlerinin HVT hücre hattında ifade edildiği bulundu. Farklı olarak, Siglec-8 sonuçları 40. döngüden sonra tespit edilemedi ve bu da negatif bir sonuç olarak kabul edildi. HVT hücre hattının immünositokimyasal incelemesinde Siglec-6 ekspresyonunun sitoplazmada orta düzeyde olduğu ortaya çıkmıştır (Skor:2). Siglec-9 ekspresyonu sitoplazmada belirgindi (Skor:3).
Sonuç: Sonuçlar Siglec-6 ve Siglec-9'un HVT hücre hattında diğer Siglec proteinlerinden daha fazla ifade edildiğini göstermiştir. İmmünositokimya sonuçları da bu bulguları desteklemektedir. Çalışmamız insan trofoblast hücrelerinde Siglec-9 ekspresyonunu gösteren ilk çalışmadır.

Keywords

Siglecs, insan villöz trafoblast, sialik asit

Project Number

2022HZDP003

Expression of sialic acid binding receptors (siglecs) in human trophoblast cell line

Abstract

Purpose: Cell surface proteins known as Siglecs exhibit a specific affinity for sialic acid. Primarily located on the surface of immune cells, Siglecs belong to the subset of lectins called I-type lectins. Siglecs have important roles in maternal-fetal immune tolerance. We aimed to analyze the expression of Siglecs on Human Villous Trophoblasts (HVT) cells.
Materials and methods: Total RNA was extracted from the HVT cell line, cDNA was synthesized, and real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR) was performed the determine the changes in Siglec -3, -5, -6, -7, -8, -9, -10, -11, and -16 mRNA levels. In addition, Siglec levels were assessed by using immunohistochemical staining. Immunoreactivity against Siglec-6 and Siglec-9 was evaluated separately according to the intensity of brown color.
Results: Expression levels of Siglec genes by qRT-PCR and melting curve analyses were performed using RNA extracted from the HVT cell line. Siglec -3, -5, -6, -7, -9, -10, -11, and -16 genes were found to be expressed in the HVT cell line. Differently, Siglec-8 results were undetected after cycle 40, which was considered a negative result. The immunocytochemical examination of the HVT cell line revealed that Siglec-6 expression was moderate in the cytoplasm (Score:2). Siglec-9 expression was prominent in the cytoplasm (Score:3).
Conclusion: The results showed that Siglec-6 and Siglec-9 were expressed more than other Siglec proteins in the human trophoblast cells. Immunocytochemistry results also support these findings. Our study is the first to show Siglec-9 expression in human trophoblast cell.

Keywords

Siglecs, human villous trophoblasts, sialic acid

Supporting Institution

This work was supported by The Pamukkale University Scientific Research Projects Coordination Unit

Project Number

2022HZDP003

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