Cholesterol and phospholipid levels of washed and percoll gradient centrifuged mouse sperm: Presence of lipids possessing inhibitory effects on sperm motility

Abstract

Levels of DNA, cholesterol, and phospholipids of mouse caudal epididymal and vas deferens sperm that were processed through simple washing and Percoll gradient centrifugation were measured. The DNA and cholesterol contents of washed sperm and Percoll gradient centrifuged (PGC) sperm (DNA = 3.6 ± 0.3 pg/sperm and 3.4 ± 0.3 pg/sperm, respectively; cholesterol = 0.219 ± 0.057 nmole/μg DNA and 0.224 ± 0.030 nmole/μg DNA, respectively, for washed and PGC sperm) were not significantly different from each other; however, the phospholipid level of PGC sperm was only one half of that of washed sperm (0.315 ± 0.071 nmole/μg DNA versus 0.720 ± 0.075 nmole/μg DNA, respectively). The presence of 0.3% bovine serum albumin (BSA) in the culture medium used in sperm washing did not change the cholesterol and phospholipid contents of washed sperm. Similarly, the cholesterol and phospholipid levels of washed sperm and PGC sperm that were further incubated in BSA‐containing medium for 30 min remained the same. Interestingly, substantial amounts of lipids, as determined by the cholesterol and phospholipid levels, were released into the supernatants of the sperm washes, and sperm needed to be washed at least twice to ensure their stable levels of cholesterol and phospholipids. The lipid mixture in the first sperm wash supernatant was shown to have inhibitory effects on PGC sperm motility. © 1996 Wiley‐Liss, Inc.

Lipidomic Profiling of Sinus Mucosa from Patients with Chronic Rhinosinusitis

“…The remainder of each homogenate and the matched human blood samples (990 uL each) were then subjected to a modified Bligh–Dyer lipid extraction . To each, 1237.5 μL of chloroform containing 300 μM BHT (CHCl 3 +BHT) and 2475 μL methanol containing 300 μM BHT (MeOH+BHT; chloroform:methanol:water ratio of 1:2:0.8, v/v/v) was added.…”

“…The remainder of each homogenate and the matched human blood samples (990 uL each) were then subjected to a modified Bligh-Dyer lipid extraction. [9][10][11][12] To each, 1237.5 μL of chloroform containing 300 μM BHT (CHCl 3 +BHT) and 2475 μL methanol containing 300 μM BHT (MeOH+BHT; chloroform: methanol: water ratio of 1: 2: 0.8, v/v/v) was added. Samples were mixed thoroughly, and after 30 minutes at 27°C, additional CHCl 3 +BHT (1237.5 uL) and dH 2 O+BHT (1237.5 uL) were added (final chloroform: methanol: water ratio of 1: 1: 0.9, v/v/v).…”

Peningkatan Kualitas Spermatozoa Pada Proses Pemisahan Spermatozoa Dengan Sentrifugasi Gradien Densitas Percoll Melalui Pemberian Fosfolipid

“…Faktor lain yang menyebabkan penurunan kualitas spermatozoa setelah proses SGDP diduga terjadi akibat pengaruh kimiawi dan mekanik langsung (fisik) gaya sentrifugasi seperti gesekan permukaan membran dengan partikel percoll atau dinding tabung. Beberapa peneliti melaporkan adanya gesekan/benturan mekanik yang terjadi antara permukaan spermatozoa dengan partikel percoll maupun dinding tabung selama proses SGDP dapat menyebabkan deformasi matriks ekstraseluler termasuk perubahan komposisi lipid membran yang penting dalam mempertahankan fluiditas membran spermatozoa (26,27). Makin lama gesekan mekanik yang terjadi pada membran spermatozoa pada proses SGDP, maka makin besar pula kerusakan integritas membran plasma yang terjadi pada spermatozoa.…”

“…Makin lama gesekan mekanik yang terjadi pada membran spermatozoa pada proses SGDP, maka makin besar pula kerusakan integritas membran plasma yang terjadi pada spermatozoa. Makin tinggi tingkat kerusakan membran plasma spermatozoa akan menyebabkan persentase spermatozoa yang memiliki membran plasma utuh makin menjadi menurun (26). Kolesterol dan fosfolipid merupakan komponen lipid membran spermatozoa yang sangat penting dalam mempertahankan integritas membran, namun sangat sensitif terhadap faktor-faktor ekstrinsik seperti suhu, dan polaritas pelarut.…”

“…Hammadeh et al, (2001) menyatakan bahwa penggunaan kuning telur dapat melindungi akrosom spermatozoa (26). Hirai et al, (1997) konsentrasi kuning telur dalam pengencer tris pada semen sapi meningkatkan persentase yang motil (14).…”