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PubMed1996

Cumulative experience of percutaneous epididymal sperm aspiration (PESA) with intracytoplasmic sperm injection.

OBJECTIVE: Our objective was to evaluate the recovery rate of spermatozoa from the epididymis using a percutaneous aspiration technique and to assess the fertilisation rate after intracytoplasmic sperm injection. MATERIALS AND METHODS: Fifty-four patients with azoospermia had a total of 59 cycles at IVF with intracytoplasmic sperm injection (ICSI). The cause of the azoospermia was failed vasectomy reversal in 23 cases, congenital absence of the vas in 22 cases, partial testicular failure in 5 cases, and retrograde ejaculation in 2 cases, while the remaining 2 patients had erectile disorders. RESULTS: A total of 741 oocytes was collected and 521 metaphase II oocytes were subsequently microinjected. Normal fertilisation occurred in 274 oocytes (52.6%), and of these, 234 cleaved (85.4%). In 54 cycles, embryo transfer of more than one embryo occurred (91.5%) and a total of 155 embryos was replaced. The pregnancy rate was 30.5% per cycle and 33.3% per embryo transfer. The implantation rate was 14.2%; failure of fertilisation occurred in two cycles, while in three other cycles the embryos did not cleave. CONCLUSIONS: Percutaneous epididymal sperm aspiration can be used successfully to retrieve sperm in men with azoospermia due to obstructive, or nonobstructive, disorders. The technique is simple, cost-effective, and associated with fewer complications than an open microsurgical operation.

Adult↗
PubMed1980

Heat activation of rat epididymal fat tissue acetyl-coa carboxylase is due to dephosphorylation by its endogenous phosphatase.

Acetyl-CoA carboxylase from rat epididymal fat tissue is activated by incubation at 30 C in the absence of citrate or metal ions. This activation is accompanied by a corresponding loss of 32P from the labeled enzyme, and it is not blocked by the heat-stable phosphorylase phosphatase inhibitor proteins from rabbit muscle. We have succeeded in separating an activity which activates and dephosphorylates acetyl-CoA carboxylase from the carboxylase using polyethylene glycol-6000. These results suggest that the temperature-dependent activation of acetyl-CoA carboxylase in crude or partially purified preparations results from dephosphorylation of the carboxylase by bound phosphatase.

Acetyl-CoA Carboxylase↗
PubMed1983

Metabolism of erucic acid in adipocytes isolated from rat epididymal fat.

The metabolism of [14-14C]erucic acid and [U-14C]palmitic acid has been investigated in adipocytes isolated from rat epididymal fat. The rate of acylation of [14C]erucic acid in cellular lipids and oxidation to CO2 and acid-soluble activity was ca. 1/3 of the rate with [14C]palmitic acid as substrate. A maximal incorporation of fatty acids in triacylglycerol was found at a fatty acid concentration of 0.8 mM in the medium, both with [14C]erucic acid and [14C]palmitic acid as substrate. Glucose added to the medium increased the esterification and decreased the oxidation of both fatty acids. No significant chain-shortening of [14C]erucic acid to shorter monoenes was identified in the fat cells. Increasing concentrations of unlabeled palmitic acid in the incubation medium markedly inhibited the esterification of [14C]erucic acid, whereas unlabeled erucic acid had little effect on the rate of esterification of [14C]palmitic acid.

Adipose Tissue↗
PubMed1992

Occurrence of wax esters and 1-O-alkyl-2,3-diacylglycerols in goat epididymal sperm plasma membrane.

Two unusual lipid classes were detected by thin-layer chromatography in the neutral lipids derived from goat cauda-epididymal sperm plasma membrane. The lipids were identified as wax esters and 1-O-alkyl-2,3-diacylglycerols based on chromatographic properties, identity of their hydrolysis products, and infrared/1H nuclear magnetic resonance spectral evidence. The membrane contained ca. 3 and 5 micrograms/mg protein of wax esters and alkyldiacylglycerols, respectively. The relative proportions of wax esters and alkyldiacylglycerols in the total neutral lipids were 1.5% and 2.4%, respectively. The lipids contained fatty acids with chain lengths of C14 to C22. The major fatty acids of the wax esters were 14:0, 16:0, 16:1 omega 7, 18:0 and 18:1 omega 9. The fatty acids in alkyldiacylglycerol were 16:0, 18:0, 22:5 omega 3 and 22:6 omega 3. Alkyldiacylglycerol was particularly rich in docosahexaenoic acid (22:6 omega 3) representing 30% of the total fatty acids. The alcohols of wax ester were all saturated with C20-C29 carbon chains. The deacylated products derived from alkyldiacylglycerols were identified as hexadecyl, octadecyl and octadec-9'-enyl glycerol ethers.

Animals↗
PubMed1983

Establishment of a preadipocyte cell line from the epididymal fat pad of the lean C57 BL/6J mouse--long term effects of insulin and triiodothyronine on adipose conversion.

A clonal cell line has been established from the epididymal fat pad of the C57 BL/6J +/? mouse. This line, designated HGFu, is aneuploid and exhibits both morphological and biochemical properties characteristic of mature adipocytes. Adipose conversion begins after confluence and is accompanied by (a) an early emergence of lipoprotein lipase, (b) an increase in the incorporation of [14C]acetate into lipids and in the activities of acid:CoA ligase and glycerol-3-phosphate dehydrogenase, (c) a 27- to 35-fold increase in the average triglyceride content per cell. In the presence of a beta-agonist (isoproterenol) a full lipolytic response (measured by fatty acid release) is observed with differentiated cells, whereas the responsiveness examined by cyclic AMP (cAMP) production is present both in undifferentiated and differentiated cells. Adipose conversion, estimated by activities of enzyme markers, is accelerated by the continuous presence in the culture medium of insulin and triiodothyronine both within their physiological range of concentrations, whereas insulin at supraphysiological concentrations shows a growth promoting activity. The concentrations of insulin and triiodothyronine required for half-maximal lipogenic effects are in agreement with the Kd values of their respective high affinity binding sites present in HGFu cells. The HGFu cell line seems to be a useful model for the study on a long term basis of the mechanisms of action both of insulin and triiodothyronine. Moreover it will make it possible to realize comparative studies between clonal lines established from the lean adult mouse (HGFu line) and from the genetically obese adult mouse (Ob17 line).

Adipose Tissue↗
PubMed1985

Sonography of epididymal cystadenoma.

Noninflammatory, epididymal masses are unusual. A patient with von Hippel-Lindau disease and sonographically demonstrated cystadenoma of the epididymis is presented.

Adult↗
PubMed2000

Characterization of Lonidamine and AF2785 blockade of the cyclic AMP-activated chloride current in rat epididymal cells.

It has been shown previously that the antifertility agents Lonidamine and its analogue AF2785, [1-(2,4-dichlorobenzyl)-indazole-3-acrylic acid] are potent inhibitors of the cAMP-activated chloride channel (CFTR) in rat epididymal cells. In this study, we further characterized the blocking actions of these two compounds and compared them with the known chloride channel blocker diphenylamine-2-carboxylate (DPC). Results show that the order of potency in blocking the cAMP-activated current is AF2785 > Lonidamine > DPC. All three compounds shared similar blocking characteristics. Firstly, their blockade of the current exhibited voltage dependence; all three agents blocked the current more markedly at negative than at positive membrane potentials. Secondly, they blocked the channels from the outside of the cell. Thirdly, their blocking efficacies were maximal at low extracellular pH. Lastly, the time course of the block by AF2785 and DPC appeared to be more rapid than that of Lonidamine. It is hoped that further studies with other indazole compounds will add knowledge to the physiology and pharmacology of CFTR in the epididymis. Such information will be of great importance to our quest for novel male contraceptives.

Animals↗
PubMed1998

Immunocytochemical alterations in the intra-acrosomal antigen MN7 during epididymal maturation of guinea pig spermatozoa.

We have previously shown that a 90-kDa intra-acrosomal antigen, MN7, is restricted to the anterior acrosomal region of mouse, rat, and hamster spermatozoa. The present study has examined the localization and the behavior of MN7 during sperm maturation in the epididymis of the guinea pig by immunoelectron microscopy. MN7 showed not only a specific localization in the apical segment of the guinea pig sperm acrosome, but also a distinct alteration during maturation, as follows. MN7 was exclusively found both at the dorsal matrix and on the outer acrosome membrane (OAM)/matrix-associated materials in the apical segment. MN7 was initially distributed throughout the electron-lucent dorsal matrix in immature sperm but, during maturation, became more restricted to the spherical bodies within the electron-lucent area. MN7 on OAM/matrix-associated materials was first distributed along the ventral margin and the small area posterior to the dorsal matrix but, during maturation, disappeared from the ventral margin and became restricted to the dorsal region. These results indicate that MN7 is a good tool for studying the stepwise maturation of epididymal spermatozoa.

Acrosome↗
PubMed2001

Predictors of success in microsurgical correction of vasal and epididymal obstruction.

Patients who inquire about microsurgical correction of vasal and epididymal obstructions should be informed about the postoperative rates of both return of sperm to the semen and pregnancy. They also should be informed about the possibility of a successful outcome, depending on their individual circumstances. The various preoperative, intraoperative, and postoperative factors that influence success rates are discussed in this article.

Epididymis↗
PubMed1988

Calmodulin-stimulated cyclic nucleotide phosphodiesterases in plasma membranes of bovine epididymal spermatozoa.

Cyclic nucleotide phosphodiesterase in the plasma membranes of bovine epididymal spermatozoa was stimulated by added Ca2+ and calmodulin. The rate of hydrolysis and responsiveness toward calmodulin was greater for cAMP than for cGMP. The kinetic analysis of the activity revealed two forms of phosphodiesterase with apparent Km values of 7.5 and 95 microM for cAMP. Calmodulin stimulated both of the activities by increasing the Vmax without affecting the Km's. The activity response with respect to Ca2+ concentration appears to be biphasic in both the absence and presence of added calmodulin. Trifluoperazine inhibited the Ca2+- and calmodulin-sensitive enzyme activity in a dose-dependent manner. The calmodulin-stimulated phosphodiesterase activity in the sperm plasma membranes can be solubilized and absorbed to a Calmodulin-Sepharose affinity column in the presence of Ca2+.

3',5'-Cyclic-AMP Phosphodiesterases↗
PubMed1987

Regulation of oxidative phosphorylation in mitochondria of epididymal bull spermatozoa.

The regulation of oxidative phosphorylation was studied with digitonin-treated epididymal bull spermatozoa in which mitochondria are directly accessible to low molecular compounds in the extracellular medium. Due to the high extramitochondrial ATPase activity in this cell preparation, it was possible to stimulate respiration to a small extent only by added hexokinase in the presence of glucose and adenine nucleotides. Added pyruvate kinase plus phosphoenol pyruvate, however, strongly suppressed the respiration. Under these conditions, the respiration was found to depend on the extramitochondrial [ATP]/[ADP] ratio in the range of 1-100. The contribution of the adenine nucleotide translocator to this dependence was determined by titration with the irreversible inhibitor carboxyatractyloside in the presence of ADP. Using lactate plus malate as substrate, the active state respiration was controlled to about 30% by the translocator, whereas 12 and 4% were determined in the presence of L-glycerol-3-phosphate and malate alone, respectively. In order to compare the results with those for intact cells, the adenine nucleotide patterns were determined in intact and digitonin-treated spermatozoa under conditions of controlled respiration in the presence of vanadate and carboxyatractyloside, respectively. About 21% of total cellular adenine nucleotides were found in digitonin-treated cells representing the mitochondrial compartment. While allowing for the intramitochondrial amount of adenine nucleotides, the cytosolic [ATP]/[ADP] ratio was estimated to be 6-times higher than the mitochondrial ratio in intact cells. It is concluded from the data presented that the principal mechanism by which oxidative phosphorylation in sperm mitochondria is regulated via the extramitochondrial [ATP]/[ADP] ratio is the same as that demonstrated for other isolated mitochondria.

Adenine Nucleotides↗
PubMed1985

Changes in the lipid content of boar sperm plasma membranes during epididymal maturation.

Plasma membranes of boar sperm from caput, corpus and cauda of the epididymis were purified by differential- and sucrose-density equilibrium centrifugation and were found to yield a single band at a density of 1.13 g/cm3. This fraction was enriched in acid and alkaline phosphatase, 5'-nucleotidase and (Na+ + K+)-ATPase activities, whereas it contained minimal amounts of hyaluronidase and N-acetylglucosaminidase and no succinic acid dehydrogenase activities. The plasma membrane of caput, corpus and cauda sperm had the same phospholipid/protein and cholesterol/phospholipid ratios but yielded different amounts of protein and individual lipid classes. Several changes in the plasma membrane were observed during transit of sperm through the epididymis. Within the phospholipid class a decrease in the percentage of phosphatidylethanolamine, phosphatidylserine and phosphatidylinositol was detected accompanied by an increase in amount of phosphatidylcholine, sphingomyelin and polyphosphoinositides. In the other lipid classes there was a decrease in the amount of free fatty acid and the major glycolipid. The amount of cholesterol decreased, while the amount of desmosterol and cholesterol sulfate increased. There was an increase in the amount of diacylglycerol. In addition, the changes in the fatty acid composition of the total membrane lipid and each phospholipid were determined. The above changes in the lipid composition of the plasma membrane during epididymal maturation may help to explain the decreased resistance to cold shock and changes in membrane fluidity of sperm during transit in the epididymis.

Aging↗
PubMed1990

Fatty acid binding protein (FABP) modulates prostaglandin E binding to rat epididymal adipocyte membrane similarly to albumin.

Albumin enhances prostaglandin E2 (PGE2) binding to isolated epididymal adipocyte membrane and also binds PGE2 with low affinity. On the other hand, S-100, ovalbumin and albumin-stearate failed to bind PGE2, as shown by ultrafiltration, and also failed to enhance PGE2 binding to the isolated adipocyte membranes. These results suggested that albumin enhances PGE2 binding possibly by serving as a carrier for the prostaglandin molecules. 3 mM warfarin or 1 mM phenylbutazone inhibited PGE2 binding to albumin by 70% and 95%, respectively, but both drugs failed to affect the enhancement of PGE2 binding to the isolated adipocyte membrane in the presence of albumin. These results exclude the possibility that PGE2 bound to albumin is more accessible to the prostaglandin receptor than free PGE2 in solution. Finally it is shown that fatty acid binding protein (FABP), a cytosolic protein which binds specifically PGE1 but not PGE2, enhances PGE1 and PGE2 binding to isolated adipocyte membranes similarly to albumin. The physiological implications of these findings are discussed.

Adipose Tissue↗
PubMed1980

Structure and metabolism of phospholipids in bovine epididymal spermatozoa.

Lipid analysis of bovine epididymal spermatozoa showed relatively large amounts of alkylacyl- and alk-1-enylacylglycerols in their choline and ethanol-amine phospholipids and alkylacylglycerol as the major constituent of a glycolipid tentatively identified as a monogalactosyl sulfate. The ether lipids exhibited remarkably simple molecular structures, i.e., the phospholipids had only 16:0 as alkyl and alk-1-enyl groups and their constituent fatty acids were almost exclusively 22:5 (n-6) and 22:6 (n-3). The glycolipid had mainly 16:0 as both alkyl and acyl moieties. In contrast, the diacyl choline and ethanolamine phosphoglycerides exhibited a much more complex fatty acid composition. 1,2-Diacylglycerols were the major nonpolar glycerolipid class and their acyl groups consisted almost exclusively of 14:0, 16:0 and 18:0. Labeled glycerol and dihydroxyacetone added to the incubation medium were readily incorporated into sperm lipids under both aerobic and anaerobic conditions. In each case, diacylcholine phosphoglycerides, diacylglycerols and phosphatidic acid were the major labeled lipids. Distribution of label among the molecular species of diacylglycerols and choline phosphoglycerides resembled somewhat their natural abundance. No radioactivity was found in alkylacyl or alk-1-enylacyl glycerolipids. The ether lipids may provide stable structural components of sperm membrane while the diacyl analogs undergo degradation and resynthesis.

Animals↗
PubMed1984

Function of calmodulin in mammalian sperm: presence of a calmodulin-dependent cyclic nucleotide phosphodiesterase associated with demembranated rat caudal epididymal sperm.

A calmodulin dependent cyclic nucleotide phosphodiesterase is associated with the head and tailpieces of demembranated rat caudal epididymal sperm. The phosphodiesterase was stimulated two-fold in the presence of Ca2+, while the simultaneous addition of Ca2+ and calmodulin resulted in a four-fold increase in activity. Ca2+ stimulation was abolished if demembranated sperm were extracted with EGTA and was recovered upon the addition of exogenous calmodulin. Micromolar levels of Ca2+ were required for full stimulation. Trifluoperazine inhibited the Ca2+ stimulated enzyme in a dose dependent manner (ID50 = 50 microM) but had no effect on the basal phosphodiesterase activity.

3',5'-Cyclic-AMP Phosphodiesterases↗
PubMed1986

Biochemical studies on proacrosin and acrosin from epididymal boar spermatozoa: in vitro translation of boar testicular proacrosin mRNA.

An inactive form of acrosin was extracted from epididymal boar spermatozoa utilizing acid pH conditions. When subjected to activation in alkaline environment, this form turns into an enzymatically active species, which exhibits close-related electrophoretic characteristics. Both the precursor and the activated species, when incubated in the presence of thermolysin, give rise to two fastly moving acrosin molecular forms. In order to establish the nature of the true acrosin zymogen, we isolated poly(A+)-RNA from boar testicles, performed its translation in vitro in the presence of [35S]-methionine and reticulocyte lysate, immunoprecipitated the translation products with anti-boar acrosin antibody, and analyzed them by SDS-polyacrylamide gel electrophoresis and autoradiography. A single translation product of molecular weight 55,000 was detected. It is concluded that the polypeptide chain of the boar zymogen is of 55,000; increases in molecular weight are due to post-translational modifications, like glycosylation.

Acrosin↗
PubMed1989

The membrane potential changes polarity during capacitation of murine epididymal sperm.

The membrane potential in murine epididymal sperm was determined with a voltage-sensitive, fluorescent probe. In freshly collected sperm, the potential was inside-negative, viz., -13 mV, and was associated with an intracellular K+ concentration of about 122 mM. Following incubation of sperm in a medium capable of sustaining capacitation and fertilization efficacy, the potential became gradually positive. An inside-positive potential, +24 mV, was obtained after 40 min of incubation, concomitant with an intracellular K+ concentration of approximately 30 mM. At this time, about 70 percent of sperm had capacitated. An inside-positive membrane potential may play a role in facilitating the acrosome reaction.

Animals↗
PubMed1989

The major maturation glycoprotein found on rat cauda epididymal sperm surface is linked to the membrane via phosphatidylinositol.

The experiments reported here further characterize a approximately 26[3H] kD cell surface glycoprotein that can be detected on rat cauda epididymal sperm using the galactose oxidase/NaB[3H]4 technique (1). When labeled sperm are treated with PI-PLC the 26[3H] kD is completely released from the cell. The released molecule can be recovered undegraded from incubation supernatant. Release by PI-PLC converts the hydrophobic, membrane-anchored form into a hydrophilic molecule as assessed by partition studies using Triton X114. Isoelectric focusing studies using both untreated (control) and PI-PLC treated samples shows that there is charge heterogeneity with two major peaks at pls of approximately 5.0 and approximately 4.5. We also show for the first time that the molecule persists on ejaculated cells.

Animals↗