[Changes in the membrane potential of the rabbit spermatozoon during epididymal maturation].
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Biomedical subjects
Publications and source records attributed to L Calzada.
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3H-sialyl residues transfer by subcellular epididymal fractions [plasma membrane (PM), microsomes (MI), and mitochondria (MT)] to both endogenous and exogenous acceptors was studied. Their fraction purity was valued (5' nucleotidase and glucose 6-phosphatase activities). The glycoprotein or glycolipid N-acetyl neuraminyl transferase activity in microsomes were 5 times higher in caput than in cauda (624 to 125 pmoles/30 min/mg protein). PM activity also was higher in caput. In MT fraction, the activity was smaller. Two mechanisms related to spermatozoa glycoprotein changes during maturation are proposed: secretion to the lumen of molecules previously glycosilated in microsomes and transialylation to spermatozoa from membranal ectoenzymes localized on the surface of the epididymal epithelium.
Normal and pathological semen were studied with regard to cholesterol and phospholipid content of sperm cells and seminal plasma. Spermatozoa from pathologic semen have similar concentrations of phospholipid-phosphorous and significantly higher cholesterol concentration than spermatozoa from normal semen. However, only oligoasthenospermic spermatozoa showed a significantly higher cholesterol/phospholipid ratio. Azoospermic seminal plasma showed the lowest values of both cholesterol and phospholipids, but the ratio of cholesterol to phospholipids was equal to that in normal spermatozoa. No significant difference was found in the cholesterol concentration of seminal plasma from oligoasthenospermic, asthenospermic, and normospermic subjects and only asthenospermic plasma showed a significantly lower concentration of this compound. Cholesterol and phospholipid exchange between sperm cells and seminal plasma was shown by the striking correlation between the lipid composition of seminal plasma with that of sperm cells.
The presence of ATPase activity was demonstrated in isolated nuclei of human spermatozoa by high resolution cytochemical methods. The Wachstein and Meisell technique as modified by Marchesi and Palade was used. ATPase activity was identified as dense and irregularly distributed granules confined to the exposed surface of spermatozoa nuclei. Within the nucleus the reaction product appeared as electron dense precipitates randomly distributed. Control experiments were negative. Deposits of lead phosphate specifically restricted to the exposed surface of nuclei were interpreted as an indication of a glucose-6-phosphatase and/or phosphohydrolase activity. Whether this activity is located in remnants of the inner leaflet of the nuclear envelope is not known. The presence of the enzyme activity within the nucleus is thought to be related to aerobic ATP synthesis previously suggested. If so, this function may be involved in establishing and/or maintaining the highly complex structural organization of spermatozoa nuclei.
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The effects on sperm metabolism of subagglutinating concentrations of infertile female sera, containing antisperm antibody, were studied. Sera were selected by their differential agglutinating or immobilizing properties and by high titers of activity. Only the immobilizing antibodies were able to decrease sperm viability. Basal and succinate depending oxygen uptake, thus as progressive motility was decreased by sera producing mixed agglutination and more drastically by that producing immobilization independent of the presence of complement. The sera with head-to-head agglutination properties induced an increase in the above mentioned physiologic parameters. Tip of tail-to-tip of tail agglutinating sera having no effect on oxygen uptake were able to induce a significant increase in progressive motility. Conversion of 14C-Glucose to 14CO2 was decreased by sera that induced tip of tail-to-tip of tail agglutination and complement-independent immobilization. The sera which induced head-to=head agglutination and immobilization on the presence of complement, were without effect. None of the tested sera showed any acitivity over the conversion of 14C-Glucose to 14C-Lactate.
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A 24-hour fast was performed in 28 normal children-17 boys and 11 girls, 2 to 17 years of age. After the fast, blood was drawn for blood sugar, plasma growth hormone and cortisol, serum free fatty acids and alanine measurements. Blood sugar values ranged between 30 and 77 mg/dl and were significantly correlated to age (R = 0.68, P less than 0.001). Plasma cortisol (R = 0.73, P less than 0.001), GH (R = 0.57, P = 0.01), and FFA (R = 0.76, P less than 0.001) were negatively correlated to age. Serum alanine fasting values ranged between 10 and 36 micrometer/dl and were significantly correlated to age (R = 0.86, P less than 0.001) and to blood sugar values (R = 0.54, P less than 0.01). These data demonstrate that carbohydrate regulation during fast improves with age in children, correlating with higher levels of gluconeogenic substrates and a lower rate of lipolysis.
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Estrogen receptor levels were determined in capacitated and noncapacitated sperm cells of normospermic men by the charcoal dextran method. The estrogen receptor of the nuclear matrix was released using hypertonic solutions. The distribution of the estrogen receptor was higher by 30% in capacitated spermatozoa as related to noncapacitated spermatozoa. The values of estrogen receptors in capacitated and noncapacitated spermatozoa were 91 +/- 21 fmol/10(8) sperm cells and 26 +/- 7 fmol/10(8) sperm cells, respectively. The association of estrogen receptor-nuclear matrix may be an important factor in the regulation of the transcription of the sperm genome of capacitated spermatozoa during the pronuclei stage.
Estrogen receptor levels were determined in nuclear matrix of human spermatozoa of normospermic fertile men and idiopathic infertile men. In noncapacitated sperm cells, the distribution of the estrogen receptor in normospermic fertile men was present in 50% of the cases, whereas in idiopathic infertile men was present in 19% of the cases. In capacitated sperm cells, the distribution of the estrogen receptor in normospermic fertile men was present in 80% of cases, whereas in idiopathic infertile men was present in 31% of the cases. The values of estrogen receptors in capacitated and noncapacitated sperm cells of normospermic fertile men were 91 +/- 21 and 26 +/- 7 fmol/10(8) sperm cells, respectively, whereas in capacitated and noncapacitated sperm cells of idiopathic infertile men, the estrogen receptor values were 50 +/- 17 and 22.5 +/- 9 fmol/10(8) sperm cells, respectively. The diminution of the estrogen receptor levels in the nuclear matrix could be a biochemical indicator of male factor infertility.
Estradiol and androgen receptor levels were determined in benign prostatic hypertrophy tissue by means of dextran-coated charcoal method. 80% of the cases contained estrogen receptor, whereas 85% of cases contained the androgen receptor. In 50 year old patients, nuclear estradiol and androgen receptors were 48% and 34% higher than the cytosolic values, respectively, whereas in 70 year old patients, the levels of nuclear estradiol and androgen receptors were 6% and 57% higher than the cytosolic values, respectively. Nuclear androgen receptor in 50 year old patients was 34% higher than the estradiol receptor, and in 70 year old patients, androgen receptor was 23% higher than the estradiol receptor. Cytosolic values of estrogen receptor in 50 year old patients were diminished 50%, whereas in 70 year old patients, the ER value was increased 40% with regard to androgen receptor. On the other hand, estradiol receptor values measured in microsomal fractions, and were higher when compared to androgen receptor levels (mean percentage +/- 50%). These findings suggest that the biological effects due to the subcellular distribution of AR or ER and that their microsomal distribution would constitute a reserve mechanism and activation of receptors for the continued growth of benign prostatic hypertrophy.
Six microsomal population of estradiol and androgen receptors have been characterized in human benign prostatic hypertrophy (BPH) and prostate cancer (PCa). Estradiol receptor (ER) and androgen receptors (AR) were extracted using 0.6 M KCL and determined by the dextran-coated charcoal method. ER and AR levels were smaller in BPH plasma membranes (PM) than in Pca cases. For functions 3, 4, 6, the ER values in PCa were 25-38% less with regard to BPH ER values. Whereas in PCa, AR values obtained in all fractions were higher when compared to BPH AR values. In benign prostatic hypertrophy and prostatic cancer, ER and AR levels were significantly higher in the nuclear fraction. In the nuclear fraction, ER and AR levels in BPH and PCa were significantly different. The subcellular distribution of AR and ER in BPH and PCa constitutes a reservation mechanism and processing a receptors for their continued growth.
The prevalence of human papillomavirus HPV-16DNA sequences in 57 penile carcinoma biopsies was examined using the polymerase chain reaction (PCR) with type specific internal probes, employing HPV consensus primers from the L1 region. The cases comprised 39 typical squamous cell carcinoma and 18 specimens with different subtype. PCR products were analyzed and HPV-16DNA was detected in a high percentage of specimens. Thirty-eight biopsies were HPV-16DNA positive. This determination was correlated with cellular differentiation and growth pattern. Our data corroborates that squamous cell carcinoma was invariably associated with HPV-16DNA.
The accumulation of the lipophilic cation radiolabeled triphenylmethylphosphonium (TPMP+) was utilized to determine the resting membrane potential across the plasma membrane (psi) of human sperm. Washed sperm were suspended and incubated in low-K+ and high-K+ medium and allowed to take up the cation to a steady state (20 min at 37 degrees C). By using this differential, the value obtained was inserted in the Nernst equation and the value yielded a psi of -69 +/- 2 mV. When the Na+ or K+ concentration is high in the medium, the accumulation of TPMP+ in the membrane sperm cells was increased or decreased, respectively, inducing hyperpolarization and depolarization of the membrane 20% and 85%, respectively. The presence of divalent cations Zn++ and Mg++ in the incubation medium both induced a hyperpolarization of 10% and 8.6%, respectively. The addition of specific reagents such as p-chloromercuribenzenosulfonate and ethylenediaminetetraacetic acid sodium salt both decreased the psi 35% and 58%, respectively. The agents acting on the sperm cell membrane, such as dithiothreitol and progesterone, both induced hyperpolarization and depolarization of the membrane 16% and 40%, respectively. The presence of propranolol and L-alpha-lysophosphatidylcholine, which affect the ionic gradients present across the plasma membrane, both induced a depolarization from 43% and 92%, respectively. Finally the psi was glucose-dependent. The result of these studies was that, by the use of agents causing hyperpolarization or depolarization, we obtained changes in the psi of -83.4 +/- 2.2 mV, until -6 +/- 0.6 mV changes of -76.8 +/- 2 mV translated across the sperm cell membrane.