# Research note: Turkey (Meleagris gallopavo) semen CD14+ macrophages – management, key features, and classification – a pilot study with optimized protocols

## TL;DR

This study isolates and characterizes turkey semen macrophages using CD14 markers to classify them into subtypes, providing a framework for future research.

## Contribution

The study introduces optimized protocols for identifying and classifying turkey semen CD14+ macrophages into Mф1 and Mф2 subpopulations.

## Key findings

- Turkey semen CD14+ macrophages are spherical, may have multiple nuclei, and contain large vacuoles.
- CD14+iNOS+ (Mф1) macrophages constitute 48.5% of CD14+ cells, while CD14+IL10+ (Mф2) macrophages make up 42.48%.
- The study establishes a framework for future investigations into the functional roles of these macrophages in turkey semen.

## Abstract

Semen macrophages (Mф) play a role in mediating innate immune responses and supporting adaptive immunity within the male reproductive tract. Knowledge about the semen Mф in avian species remains limited. This study aimed to isolate turkey semen Mф and optimize protocols for their characterization based on specific Mф markers, specifically cluster of differentiation 14 (CD14), inducible nitric oxide synthase (iNOS), and interleukin 10 (IL10), for further classification into CD14+ type 1 macrophages (CD14+iNOS+, akin CD14+Mф1) and CD14+ type 2 macrophages (CD14+IL10+, akin CD14+Mф2) subpopulations. In the current study, immunofluorescence (IF) staining and fluorescence-activated cell sorting (FACS) were employed to identify these markers, and the morphological features of CD14+Mф were examined. Our results demonstrated that turkey semen CD14+Mф are spherical, may contain one or more nuclei, and often exhibit large vacuoles. Based on FACS results, CD14+iNOS+(akin CD14+Mф1) accounted for 48.5 ± 12.45 % of CD14+ cells, whereas CD14+IL10+ (akin CD14+Mф2) represented 42.48 ± 14.62 % of them. This study provides a foundational framework for future investigations into the functional roles of turkey CD14+Mф in semen.