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C J Flickinger

Publications and source records attributed to C J Flickinger.

142 records · Page 8Linked to original sources
PubMed1969

The development of Golgi complexes and their dependence upon the nucleus inmebae.

The production of Golgi complexes was investigated in Amoeba proteus by introducing a nucleus into cells that had been enucleated for 5 days. Golgi complexes were not detected in 5 day enucleates, nor were they observed in amebae fixed 15 min after renucleation. Samples taken at longer intervals after the introduction of a nucleus exhibited an increase in the size and abundance of Golgi complexes. Small curved smooth cisternae, some of which were aligned in parallel to form small Golgi complexes, were observed 30 min after the operation. Aggregations of small Golgi complexes increased in number in amebae fixed 1 to 6 hr after renucleation. Golgi complexes of normal size were present 6 hr after the operation and became more abundant in samples fixed 12 hr, and 1, 2, and 3 days after renucleation. The possible participation of the granular endoplasmic reticulum in the development of Golgi complexes was suggested by two observations. First, the Golgi complexes in renucleates contained a dense material similar to the content of the endoplasmic reticulum in enucleates and early renucleates. Second, examples of continuity between the endoplasmic reticulum and Golgi cisternae were present in renucleates. The possibility that Golgi complexes can be produced in the absence of preexisting Golgi complexes is discussed.

Amoeba↗
PubMed1968

The effects of enucleation on the cytoplasmic membranes of Amoeba proteus.

The dependence of cytoplasmic membranes upon the nucleus was studied by examining enucleated amebae with the electron microscope at intervals up to 1 wk after enucleation. Amebae were cut into two approximately equal parts, and the fine structure of the enucleated portions was compared with that of the nucleated parts and starved whole cells which had been maintained under the same conditions. Golgi bodies were diminished in size 1 day after enucleation and were not detected in cells enucleated for more than 2 days. The endoplasmic reticulum of enucleated cells appeared to increase in amount and underwent changes in its morphology. The sparsely scattered short tubules of granular endoplasmic reticulum present in unmanipulated amebae from stock cultures were replaced in 1-3-day enucleates by long narrow cisternae. In 3-7-day enucleates, similar cisternae of granular endoplasmic reticulum encircled areas of cytoplasm partially or completely. It was estimated that in most cases hundreds of these areas encircled by two rough membranes were formed per enucleated cell. The number of ribosomes studding the surface of the endoplasmic reticulum decreased progressively with time after enucleation. In contrast, the membranes of nucleated parts and starved whole cells did not undergo these changes. The possible identification of membrane-encircled areas as cytolysomes and their mode of formation are considered. Implications of the observations regarding nuclear regulation of the form of the Golgi apparatus and the endoplasmic reticulum are discussed.

Amoeba↗
PubMed1965

The fine structure of the nuclei of Tetrahymena pyriformis throughout the cell cycle.

The fine structure of the nuclei of logarithmically growing Tetrahymena pyriformis, strain HSM, was studied at 30-minute intervals throughout the cell cycle. Organisms were selected at similar stages of cytokinesis by means of a braking pipette, incubated, fixed in OsO(4), and embedded in agar to facilitate subsequent preparation for electron microscopy. Aggregates of micronuclear chromatin underwent a decrease in density and number with a concomitant increase in size throughout interphase. There were no impressive changes in macronuclear morphology. It was found possible to estimate a cell's progress through interphase by observation of micronuclear morphology, but attempts to correlate changes in fine structure with periods of DNA synthesis were unsuccessful.

Cell Division↗
PubMedDate not supplied

Synthesis and secretion of glycoprotein by the epididymal epithelium.

The secretory activity of the middle segment of the mouse caput epididymidis was studied using 3H-fucose as a precursor to glycoprotein. Young adult male mice were injected with a concentrated solution of 3H-fucose interstitially, ie beneath the connective tissue capsule of the epididymis. Two animals were killed and prepared for light microscopic radioautography at each of six intervals between 10 minutes and 24 hours after injection. Silver grains were concentrated over the supranuclear Golgi region at 10 minutes and over the apical ends of the cells 30 minutes and 1 hour after injection. Quantitative analysis showed that luminal radioactivity increased greatly beginning with the 2-hour samples. The results indicate that the epididymal epithelium synthesizes and secretes glycoproteins, and that 1 to 2 hours are required for terminal glycosylation, intracellular transport, and release of the secretory product.

Animals↗
PubMedDate not supplied

Identification and isolation of epididymal luminal proteins of the mouse.

Proteins of mouse cauda epididymal fluid were analyzed by polyacrylamide gel electrophoresis. Fluid expressed from the cauda epididymidis and samples obtained by micropuncture of the epididymal lumen showed very similar patterns with respect to the major proteins they contained, with the exception of a small amount of serum albumin found in expressed caudal fluid. Eight prominent peptides present in both micropuncture fluid and expressed caudal epididymal fluid were selected for further study, and were designated CP 47, 42, 35, 29, 27, 25, 18, and 13 according to their mobility. Six of these were never detected in serum. Periodic acid-Schiff staining indicated that at least three were glycoproteins. The epididymal proteins were purified by preparative polyacrylamide gel electrophoresis and electroelution. Upon re-electrophoresis, the individual purified peptides comigrated with the corresponding bands in whole epididymal fluid, and no additional bands were detected, indicating that the proteins were purified to a high degree of homogeneity. Several of the mouse epididymal peptides resemble in their mobility proteins identified previously in other species, most notably the widely studied 33 Kd and 16 to 18 Kd proteins detected in the rat.

Animals↗
PubMedDate not supplied

The incidence of spermatic granulomas and their relation to testis weight after vasectomy and vasovasostomy in Lewis rats.

The occurrence of spermatic granulomas of the vas deferens was studied in Lewis rats at intervals up to 7 months after vasectomy or vasectomy followed 3 months later by vasovasostomy. The incidence of granuloma progressed with time to involve one or both tracts in 100% of vasectomized rats. In addition, the majority of animals developed new granulomas after vasovasostomy, even though fluid flow through the reconnected vas deferens was demonstrated in vitro. When individual tracts were analyzed, the weight of the testis was related to ipsilateral spermatic granuloma formation in both vasectomy and vasovasostomy groups at 3 and 4 months after initial operation. Testes were small in the absence of a granuloma but similar to those of sham-operated rats if a granuloma was present. The possible protective effect of spermatic granuloma formation on the testis is discussed.

Animals↗
PubMedDate not supplied

Temporal appearance of antisperm autoantibodies in Lewis rats following vasectomy.

An indirect enzyme-linked immunosorbent assay (ELISA) was employed to monitor antisperm autoantibodies in 16 Lewis rats for up to 36 weeks following vasectomy. This assay was capable of discriminating all prevasectomy from postvasectomy sera at a 1:16 dilution. Weekly serum samples were obtained for the first 13 weeks and bimonthly samples thereafter. Half of the animals developed a positive antisperm autoantibody response by the end of the first postoperative week. By the end of the second week, 81% of the animals had positive responses. The greatest proportion (88%) of animals having a positive response over the course of the study was found at the end of the seventh postoperative week and the highest mean absorbance value for all 16 animals was observed at this time. Only 25% of the animals had positive responses for antisperm autoantibody at the end of the 35th week of the study. These findings indicate that circulating antisperm autoantibodies arise in the Lewis rat earlier than has been generally appreciated. The time course is similar to that of antibody titers to infectious agents or arising from inoculation of rats with spermatozoa. These findings on autoantibody levels in the Lewis rat are compared with the dynamics of antisperm autoantibody formation in man.

Animals↗
PubMedDate not supplied

Postvasectomy alterations in protein synthesis and secretion in the rat caput epididymidis are not repaired after vasovasostomy.

Many men who have undergone vasectomy later request vasovasostomy. Unfortunately, significant numbers of these men remain infertile despite the reestablishment of patent ducts. This report examines the possibility that epididymal function remains compromised after vasovasostomy in the rat by examination of quantifiable, in vivo protein synthesis and secretion in the caput epididymidis. Rats were studied 30 days after vasectomy, 30 days after a vasovasostomy (which was performed 30 days after vasectomy), or after sham operations. Epididymal lumen fluids (LF) were collected by micropuncture after 3 hours' in vivo microperifusion of tubules with 35S-amino acids. Proteins were separated by 2-dimensional electrophoresis and were detected by Coomassie blue staining. Synthesized proteins in tubule extract and synthesized and secreted proteins in LF were detected by autoradiography and image analysis. Specific proteins that appeared to be affected by vasectomy-vasovasostomy were identified by internal sequence analysis. LF contained an average of 87 detectable proteins synthesized and secreted in the control caput. Nineteen of the most prominent LF proteins were selected for more focused study. The most prominent proteins were clusterin, cysteine-rich secretory protein (CRISP)-1, and epididymal retinoic acid-binding protein. Among these, CRISP-1 remained reduced in LF after vasovasostomy. Two more minor proteins that remained reduced after vasovasostomy were identified as prostaglandin D2 synthase and phosphatidylethanolamine-binding protein. All 3 of these proteins occur in the epididymides of multiple species and have been associated with sperm fertilizing capacity.

Animals↗
PubMedDate not supplied

Assignment of the gene for human intra-acrosomal protein SP-10 to the p12----q13 region of chromosome 11.

The human sperm antigen SP-10 is a testis-specific, intra-acrosomal protein associated with the membranes of the acrosomal vesicle. The molecule has been designated a "primary vaccine candidate" by a World Health Organization (WHO) Taskforce on Contraceptive Vaccines. cDNA cloning and sequencing have indicated that SP-10 is encoded by a 795-base-pair (bp) reading frame that predicts a 265-amino acid protein of 28.3 kd. In this study, we used a 634-bp fragment (bp 68 through 700, amino acids 3 through 222) of the SP-10 sequence to probe, by Southern blotting, EcoRI-digested DNA from 33 mouse/human somatic cell hybrids involving 16 unrelated human cell lines and 4 mouse cell lines. The hybrids were characterized by karyotypic analysis and by mapped enzyme markers. The presence or absence of positive human bands was scored on the blots and the percent of concordance and discordance with a specific human chromosome was determined. The DNA probe for SP-10 showed a concordance of 31 and a discordancy of 0 for human chromosome 11, mapping SP-10 unequivocally to this chromosome. The hybrid XER-7 with the 11/X translocation: 11p12 or 11p11----11qter:: Xq11----Xqter and the hybrid EXR-5CSAZ with the X/11 translocation: Xpter----Xq22::11q13----11qter localized the SP-10 gene to the p12----q13 region. The SP-10 locus has been assigned the gene symbol ACRV1 (acrosomal vesicle protein-1).

Acrosome↗
PubMedDate not supplied

Temporal appearance of antisperm antibodies during sexual maturation of rats after obstruction of the vas deferens.

The appearance of serum antisperm antibodies was studied during the maturation of rats in which the vas deferens was obstructed prepubertally. Vasa deferentia were ligated and divided bilaterally in 10-day-old Lewis rats, while control animals received a sham operation at the same age. At ages between 14 and 128 days, blood samples were assayed for antisperm antibodies using an enzyme-linked immunosorbent assay. There were no differences between obstructed and sham-operated groups in mean antisperm antibodies from 14 through 56 days. However, in 91- and 128-day-old rats mean antisperm antibody levels were much higher in obstructed than in control animals. This rise in antisperm antibodies occurred several weeks after the development of spermatids in the testes between 23 and 46 days. The presence of elevated antisperm antibodies in obstructed animals followed temporally the appearance of sperm throughout the lumen of the entire epididymis and the formation of spermatic granulomas. The results in this model suggest that consideration be given to repair of injured or obstructed vas deferens prior to puberty to forestall development of antisperm antibodies, as well as the formation of spermatic granulomas.

Animals↗
PubMedDate not supplied

Sperm autoantigens recognized by autoantibodies in developing rats following prepubertal obstruction of the vas deferens.

Sperm protein autoantigens recognized by serum antisperm autoantibodies during development following vas deferens obstruction were studied using western blot analysis. At age 10 days, rats in an obstructed group underwent bilateral ligation and division of the vas deferens, whereas animals assigned to a sham group received a sham operation. At ages 14, 21, 35, 56, 91, and 128 days, rats were sacrificed and blood samples were obtained. Sperm antigens were recognized infrequently and with low intensity by most sera obtained at ages 14 through 56 days. Subsequently, the incidence as well as the intensity of staining of antigens in blots increased at 91- and 128-day intervals in obstructed animals. The increase in antisperm antibodies correlated with the appearance of sperm throughout the epididymis at approximately 56 days postnatally. A set of autoantigens including proteins migrating at 82-78, 76-73, 68, 57, 54, 48, 44, 42, 38-42, 36, and 22 kDa were recognized with the highest frequency and intensity after puberty. A 42-kDa protein appeared to be one of the first autoantigens recognized when obstructed animals underwent sexual maturation. Although individual animals recognized different patterns of sperm autoantigens, a repertoire of 10-12 autoantigens dominated the antisperm antibody response when obstructed animals underwent sexual maturation.

Animals↗
PubMedDate not supplied

Epididymal obstruction during development results in antisperm autoantibodies at puberty in rats.

An autoimmune response to sperm occurs after vasectomy, but there is little information on whether similar reactions occur after obstruction of the male reproductive tract at other points. Male Lewis rats received bilateral ligation of the corpus epididymidis or a sham operation at age 10 days, and the subsequent systemic antisperm autoantibody responses were compared to those observed following obstruction of the vas deferens. After sexual maturation, rats with epididymal ligations had antisperm antibodies on an enzyme-linked immunosorbent assay that were significantly higher than those of sham-operated animals and did not differ from antibody levels in vasectomized rats at the same ages. Western blot analysis showed that certain sperm proteins were recognized by antisperm antibodies after both epididymal ligation and vasectomy, including the previously identified "dominant" autoantigens at 73-83, 68-72, 48, 42, and 22 kDa. On the other hand, sera from rats with epididymal ligations recognized 60 and 52 kDa proteins that were not bound by most postvasectomy sera. Conversely, 42-48 and 38-42 kDa bands were more strongly and frequently stained after vasectomy than after epididymal ligation. The results demonstrate that antisperm antibodies are produced after obstruction of the epididymis and that the magnitude of the response is comparable to that after vasal obstruction. Differences in autoantigens recognized after epididymal and vasal obstructions may reflect maturational changes in sperm components that take place during the passage of spermatozoa through the epididymis.

Animals↗