Purification of cryopreserved camel spermatozoa following protease‐based semen liquefaction by lectin‐functionalized DNA‐defrag magnetic nanoparticles

Abstract

Although incorporating proteases into sperm medium is considered the most effective procedure to eliminate camel semen viscosity, it drastically affects viability, morpho‐functional properties and, hence, fertilization potential of spermatozoa. The present work aimed at evaluating adequacy of employing magnetic nanoparticles‐based sperm purification technique for eluting impaired and apoptotic camel spermatozoa from cryopreserved semen doses following protease‐based semen liquefaction. Thirty cryopreserved semen doses (50 x 106 sperm/straw) representing the following liquefaction treatments: control (untreated), 0.1 mg/ml papain or 5 U/ml bromelain were used (n = 10 straws per treatment). Immediately after thawing (38°C for 40 s), sperm concentration of each straw within treatment was readjusted to 15 x 106 sperm/mL by dilution in PBS (37°C). Sperm physical and cytological properties were then assessed (non‐purified semen). Thereafter, each specimen was subjected to lectin‐functionalized DNA‐defrag magnetic nanoparticles sperm purification, and the same sperm traits were re‐evaluated after undergoing purification (purified semen). Sperm DNA fragmentation level within each group, prior to and after magnetic nano‐purification, was also determined by fluorescent imaging. The results showed a dramatic improvement (p < .05) in post‐thaw motility (%), viability (%), normal sperm (%), intact acrosome (%) and HOST‐reacted (%) spermatozoa in protease‐liquefied semen following sperm magnetic nano‐purification. Additionally, the highest (p < .05) DNA fragmentation level was recorded in all cryopreserved semen groups prior to purification, whereas the lowest (p < .05) was observed in the protease‐treated specimens after magnetic nano‐purification. These results indicate that protease‐based semen liquefaction prior to cryopreservation in conjunction with magnetic nano‐purification post‐thawing holds potential for reducing the proportion of damaged and dead spermatozoa, hence improving camel sperm fertilization competence.

Seminal Microbiota of Idiopathic Infertile Patients and Its Relationship With Sperm DNA Integrity

“…These bacteria might benefit from a more liquid medium, which would allow for higher mobility and would made resource availability easier, giving them a proliferative advantage. Alternatively, some of these genera might act directly on the viscosity of the medium through the production of acids or hydrolase enzymes, which would degrade seminal fluid proteins and promote their liquefaction ( Kershaw-Young et al, 2013 ; da Fonseca Junior et al, 2020 ; Rateb, 2021 ).…”

The Positive Effects of In Vivo/In Vitro Suplementation of Nanoparticles on Semen

“…Across generations, phenotypic alterations ignited by embryo manipulations are believed to persist, according to previous research (Dominiquez et al, 2020;2021). Nanomaterials used in manipulations during embryonic development may pose future health risks and safety concerns that demand attention.…”

“…As part of the apoptotic process, DNA fragmentation occurs, although this by itself does not have a major impact on the effectiveness of assisted reproduction. Cells that have begun the dying process but have not yet reached the point of ultimate collapse may still contain intact DNA since DNA fragmentation is a late indicator of apoptosis (Rateb, 2021). Loss of membrane integrity, a prelude to apoptosis , is even more intriguing.…”

“…This reduces their commercial viability since there is less high-quality germplasm available for breeding propagation. Increased oxidative stress because of the presence of dead and injured spermatozoa (25%-30%) producing ROS may be a primary cause of ejaculate rejection (Bisla et al, 2020;2021;Kumar et al, 2018;Rautela et al, 2020).…”

Biocompatible Nanomaterials as an Emerging Technology in Reproductive Health; a Focus on the Male

“…Damaged and apoptotic sperm remain attached to the magnetic beads whilst the flow through carries the better quality sperm from the column. Nanoparticles such as magnetic IONPs coated with lectins or a combination of lectins and Annexin V that bind to, and facilitate removal of, damaged spermatozoa have been applied to porcine ( Feugang et al, 2015a ; Durfey et al, 2019 ; Feugang et al, 2019 ) and camel ( Rateb, 2020 ) semen samples prior to downstream application in AI, and to human spermatozoa for subsequent use in ICSI ( Pacheco et al, 2020 ). Additionally, in a study aimed at improving the quality of spermatozoa in cattle semen samples, Farini et al (2016) used Cell-SELEX to discover ssDNA aptamers that bind to damaged spermatozoa and applied these to the surface of magnetic IONPs.…”