Immunolocalization of the telomerase‐1 component in cells of the regenerating tail,testis, and intestine of lizards

Using an antibody against a lizard telomerase‐1 component the presence of telomerase has been detected in regenerating lizard tails where numerous cells are proliferating. Immunoblots showed telomerase positive bands at 75–80 kDa in normal tissues and at 50, 75, and 90 kDa in those regenerating. Immunofluorescence and ultrastructural immunolocalization showed telomerase‐immunoreactivity in sparCe (few/diluted) mesenchymal cells of the blastema, early regenerating muscles, perichondrium of the cartilaginous tube, ependyma of the spinal cord, and in the regenerating epidermis. Clusters of gold particles were detected in condensing chromosomes of few mesenchymal and epithelial cells in the regenerating tail, but a low to undetectable labeling in interphase cells. Telomerase‐immunoreactivity was intense in the nucleus and sparCe (few/diluted) in the cytoplasm of spermatogonia and spermatocytes and drastically decreased in early spermatids where some nuclear labeling remains. Some intense immunoreactivity was seen in few cells near the basal membrane of intestinal enterocytes or in leukocytes (likely lymphocytes) of the intestine mucosa. In spermatogonia, spermatids and in enterocytes part of the nuclear labeling formed cluster of gold particles in dense areas identified as Cajal Bodies, suggesting that telomerase is a marker for these stem cells. This therefore suggests that also the sparCe (few/diluted) telomerase positive cells detected in the regenerating tail may represent sparCe (few/diluted) stem cells localized in regenerating tissues where transit amplifying cells are instead preponderant to allow for tail growth. This observation supports previous studies indicating that few stem cells are present in the stump after tail amputation and give rise to transit amplifying cells for tail regeneration. J. Morphol. 276:748–758, 2015. © 2015 Wiley Periodicals, Inc.     

Angioarchitecture of the bovine spermatic cord

We described the topography and morphometry of the testicular artery, pampiniform plexus veins, and indirect connections between them in the spermatic cord of the bull. Sixty microcorrosive casts of bovine spermatic cords were analyzed macroscopically, by stereomicroscopy, and by scanning electron microscopy. The average size of the testicles was 94.6 × 49.7 × 54.7 mm. The testicular artery formed a superiorly pointed cone‐like structure with its base fixed to the proximal part of the gonad. The artery gave off one or two branches to the head of epididymis and to the deferens duct. The pampiniform plexus originated from intra‐tunical veins. Veins of the pampiniform plexus were of smaller diameter but larger number than intra‐tunical ones. The density of the veins of the pampiniform plexus was 9.37 ± 1.07 mm^?2. The testicular vein began 90–121 mm above the superior pole of the testis. In 2.9% of specimens, the testicular vein was doubled. Numerous anastomoses among veins of pampiniform plexus were observed. Additionally, indirect anastomoses between the testicular artery and pampiniform plexus veins formed by the capillary network of the vasa vasorum of the testicular artery were visualized by scanning electron microscopy. In all cases, narrowings in the casts of the precapillary vessel were observed. We also documented the vasa vasorum of the testicular artery in bulls. The density of these vessels was 22.87 ± 11.48 mm^?2. The indirect arteriovenous connections together with the presence of circular constrictions of the lumen in precapillary vessels may play a role in testicular blood flow regulation. J. Morphol., 2011. © 2011 Wiley‐Liss, Inc.     

Rete testis fluid contains a growth factor for cultured fibroblasts

A new polypeptide mitogen has been detected at high specific activity in the rete testis fluid of rams (oRTF). The factor, which stimulates DNA synthesis in quiescent Swiss 3T3 cells, has a molecular weight of 45,000 as assessed by gel filtration through Ultrogel AcA 34. The factor is heat stable but is inactivated by proteolytic enzymes and by β-mercaptoethanol. The growth-promoting activity in oRTF does not bind to concanavalin A.     

Effects of fucoidan on diabetic rat testicular tissue

Increased reactive oxygen species (ROS) resulting from hyperglycemia and inadequate endogenous antioxidant systems are responsible for the complications of diabetes. ROS accumulate in the cell and stimulate apoptosis, which compromises sperm quality and function. We investigated the possible effects of fucoidan, a potent antioxidant with a regulatory effect on blood glucose homeostasis, on the testicular tissues of rats with experimental diabetes. Diabetes was induced by administering 40 mg/kg streptozotocin (STZ) on five consecutive days. Twenty-four Wistar albino male rats were divided into four groups: group 1, control group (CG); group 2, diabetes group (DG); group 3, early fucoidan group (EFG) treated with 50 mg/kg fucoidan after diabetes induction; group 4, late fucoidan group (LFG) treated with the same dose of fucoidan 15 days after diabetes induction. Fucoidan was administered intraperitoneally every two days for four weeks. Basement membrane thickness and Johnsen scores were higher in the DG than in the CG; no difference was found for either the EFG or LFG compared to the CG. Seminiferous tubule diameters of EFG were significantly greater than for the DG. Apoptotic tubule and apoptotic cell indexes were significantly greater in the DG and significantly less in the EFG and LFG groups compared to the CG. Early use of fucoidan in diabetic individuals may minimize damage to testicular tissue.     

Human spermatogenic cell marker proteins detected by two-dimensional electrophoresis

Highly homogeneous populations of human pachytene spetmatocytes and round spermatids have been obtained from normal adult testis using unit gravity (STA-PUT) sedimentation. Contaminating Leydig cells have been removed by density centrifugation in discontinuous Percoll gradients to yield resultant germ cell purities of 90–95% for pachytene spermatocytes and 89–96% for round spermatids. The total cellular polypeptide composition of separated human germ cells has been analyzed by two-dimensional polyacrylamide gel electrophoresis to compare 1) human and mouse pachytene spermatocytes (species specificity), 2) samples of human spermatocytes obtained from different individuals (allo specificity), and 3) pachytene spermatocytes and round spermatids from the same patients (stage specificity). Mouse and human germ cells have been found to exhibit extensive homology, but identified marker proteins limited to human spermatocytes include a group of polypeptides at p45/5.9 as well as a protein at p67/5.2. Proteins unique to mouse germ cells include component p65/5.5. Comparisons between different preparations of human pachytene spermatocytes have revealed about 90% electrophoretic homology, but some striking allotypic variations have been noted including the proteins at p45/5.9. Finally, presumptive stage-specific spermatogenic cell markers have been identified including the p45/5.9 polypeptides that are present only in human spermatocytes. Although the physiological roles of particular marker proteins have not yet been determined, the present findings indicate that purified spermatogenic cell populations may be analyzed biochemically to identify constituents important in the regulation of sperm development in man.     

Morphometric studies on mink testicular tissue

The mink, a seasonal breeder of great economic importance, shows a high incidence of male infertility. This problem has forced investigators to find methods of assaying male mink infertility. In this study, morphometric studies have been performed on testicular tissue of a total of 31 males eliminated from breeding after testicular palpation, sperm test, and estimation of serum testosterone concentrations. Males having low sperm quality or disturbed testicular development (n=24) had significantly (p<0.01) lower numbers of spermatocytes, spermatids, and freefloating luminal spermatozoa. compared with males with good sperm quality (n=7). No differences were found in the numbers of spermatogonia, Sertoli, and Leydig cells. Other morphometric parameters such as mean diameter, mean area, mean volume, percentage of area, and surface area per volume of nuclei are also presented for each cell type in the testis. It may be concluded that the sperm test is best suited for assessing fertility in mink. Severe disturbances in testicular development can be detected by testicular palpation and serum testosterone measurements.     

Comparison of androgen biosynthesis in isolated leydig cells from rat and mouse testis: incubation and superfusion studies

Production of testosterone by highly purified Leydig cells prepared from rat and mouse testes is compared. Testosterone formation is improved to a higher degree in rat (2.7-fold) than in mouse (1.7-fold) cells by collagenase treatment of the testis compared with mechanical isolation. Mouse Leydig cells respond to exogenous stimuli (choriogonadotropin, dibutyryl cyclic AMP) with 2.4-fold higher testosterone secretion than rat cells. A 1.7-fold increased conversion of androgen precursors to testosterone by mouse compared with rat Leydig cells is demonstrated in static incubations as well as in steady-state superfusion experiments and can be derived from enhanced androstenedione reduction and a less inhibitory effect of progesterone on this process in mouse Leydig cells.     

Purification,characterization and partial amino acid sequencing of a 70 kD inhibitor protein of Na+,K+-ATPase from goat testis cytosol

A protein isolated from goat testis cytosol is found to inhibit Na⁺,K⁺-ATPase from rat brain microsomes. The inhibitor has been purified by ammonium sulphate precipitation followed by hydroxyapatite column chromatography. The purified fraction appears as a single polypeptide band on 10% SDS-PAGE of approximate molecular mass of 70 kDa. The concentration at which 50% inhibition (I₅₀) occurs is in the nanomolar range. The inhibitor seems to bind Na⁺,K⁺-ATPase reversibly at ATP binding site in a competitive manner with ATP, but away from ouabain binding site. It does not affect p-nitrophenyl-phosphatase activity. The inhibitor is found to inhibit the phosphorylation step of the Na⁺,K⁺-ATPase. The enhancement of tryptophan fluorescence and changes in CD pattern suggest conformational changes of Na⁺,K⁺-ATPase on binding to the inhibitor. Amino acid sequence of the trypsinised fragments show some homology with aldehyde reductase.     

Brain and testis: more alike than previously thought?

Several strands of evidence indicate the presence of marked similarities between human brain and testis. Understanding these similarities and their implications has become a topic of interest among the scientific community. Indeed, an association of intelligence with some semen quality parameters has been reported and a relation between dysfunctions of the human brain and testis has also been evident. Numerous common molecular features are evident when these tissues are compared, which is reflected in the huge number of common proteins. At the functional level, human neurons and sperm share a number of characteristics, including the importance of the exocytotic process and the presence of similar receptors and signalling pathways. The common proteins are mainly involved in exocytosis, tissue development and neuron/brain-associated biological processes. With this analysis, we conclude that human brain and testis share several biochemical characteristics which, in addition to their involvement in the speciation process, could, at least in part, be responsible for the expression of a huge number of common proteins. Nonetheless, this is an underexplored topic, and the connection between these tissues needs to be clarified, which could help to understand the dysfunctions affecting brain and testis, as well as to develop improved therapeutic strategies.