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A rat model to study the local effects of testicular vein ligation is described. One hour after unilateral testicular vein ligation, the testicular concentration of testosterone was significantly greater (P less than 0.01) on the side that was ligated than in the contralateral testis (177.1 +/- 19.7 [SEM] ng/gm of tissue as compared with 108.8 +/- 11.8 ng/gm of tissue, respectively). This effect was not seen 1 week after the testicular vein ligation. The testosterone concentration in the ligated testis was also higher than that in sham-operated animals. These differences in testicular testosterone concentration were not associated with changes in peripheral serum testosterone levels. ligation of the testicular vein causes an acute rise in the testicular concentration of testosterone and may thus mediate changes in testicular function. Show less

Protein carboxyl-methylase (PCM), the enzyme that transfers methyl groups from S-adenosyl-methionine to free carboxyl groups on proteins, is highly localized in testes. The cellular distribution of PCM and its substrates, the methyl acceptor proteins, was investigated. Separation of testicular cells on an albumin gravity gradient revealed the preferential localization of both enzyme and substrates in spermatids. In young rats, PCM activity increases with age coincidently with germ cell maturation. Rats which are heterozygous for the Hre gene (Hre/+) are infertile as a result of germ cell depletion. In these animals, testicular PCM specific activity and total activity were, respectively, 4--6 and 40--50 times lower than in normal testes. Enzyme activity in testes from animals with x-ray-induced germ cell depletion was also very low. These observations suggest that PCM is located in germ cells. Show less

In the light of earlier work [1] which demonstrated the presence of a large number of myosin heavy chain (MHC) transcripts in chick myoblasts prior to cell fusion and the burst of MHC synthesis it was of great interest to determine the subcellular localization of the still inactive transcripts. It has been determined in differentiating muscle cells in culture. Two populations of cells were examined -- monucleated myoblasts just prior to cell fusion and myotubes where at least 80% of the cells were fused. Utilizing a myosin complementary DNA (cDNA) probe [2] it is observed that just prior to cell fusion, when the "burst" of myosin synthesis has not yet occurred, the vast majority of cytoplasmic myosin mRNA transcripts are found in a stored messenger RNA protein complex with a minimal amount found in the heavy polysome fraction. In differentiated myotube cultures, when myosin synthesis is progressing at a high rate, the reverse is found, i.e, the amount of stored myosin messenger RNA (mRNA) is minimal while the largest amount of myosin mRNA transcripts are localized in the polysome fraction. The number of total cytoplasmic myosin transcripts is found to decrease after cell fusion at a time when myosin synthesis is maximal suggesting that the efficiency of translation of myosin mRNA increases during terminal differentiation. Show less

Double lip, an accessory fold of redundant mucous membrane inside the vermilion border, is differentiated from other chronic enlargements of the lip, and treatment for this congenital anomaly is outlined. Show less

Administration of FSH antiserum to adult rats for 14 or 30 days had no or little effect on body, testis or accessory sex gland weights, androgen-binding protein, testosterone levels, germ cell numbers or fertility, thus indicating a relative insensitivity of the testis to withdrawal of FSH. Unlike immature rats, therefore, which do require FSH to initiate spermatogenesis, adult rats do not need this hormone to maintain spermatogenesis. Show less

Monochromatic targets presented at 30 degrees excentricity on orange, magenta and blue backgrouds are used. A small monochromatic light, 476 nm on orange, 551 nm on magenta and 621 nm on blue, is flashed at 3 cps-1 on the centre of the targets. The size of the targets is varied and their luminance adjusted using neutral filters until the flashing light is just not visible. This method allows the study of chromatic mechanism sensitivity and of retinal interactions (summation and inhibition). Some observations in normal as well as in pathological conditions are presented. Show less

Different types of human germ cells show unusual features of the nuclear envelope. Spermatogonial nuclei demonstrate two kinds of modifications. The first one is a series of intranuclear flattened cisterns, parallel to each other and to the inner aspect of the nuclear envelope. The second one is a nuclear envelope protrusion into the cytoplasm occupied by a double membrane-limited vesicle. Pores are found on the membrane of the vesicle facing the interior of the nucleus. In spermatocytes the nuclear pores are concentrated over certain areas and completely absent from others. In the regions where they are absent a single cytoplasmic cistern of rough endoplasmic reticulum is closely apposed to the outer membrane of the nuclear envelope. Early modifications of the nuclear surface appear in spermatids before the attachment of the acrosomic vesicle and may indicate an active role of the nuclear envelope in the morphogenesis of the acrosome. In round spermatids nuclear pores are absent from the area which is first related to the Golgi and later covered by the acrosomal cap. Single or multiple layers of cytoplasmic annulate lamellae are closely associated with the nuclear envelope over the pore rich areas. Frequently there are intranuclear accumulations of dense material adjacent to the annulate lamellae-nuclear pore complex. The chromatoid body is usually present on the cytoplasmic side of this complex. In the elongating spermatids most annulate lamellae are free in the cytoplasm, often in relation with Golgi and chromatoid body remnants near the axial filament. Few stacks of annulate lamellae are noted adjacent to the pore rich nuclear regions. It is suggested that the described modifications are related to an active nuclear-cytoplasmic interaction. Show less

A clone panel containing various segments of human chromosome 11 has been selected and use for regional assignment of the gene for human lysosomal acid phosphatase (ACP2) to the short arm of chromosome 11, in the region 11p11 leads to 11p12. Further evidence has also been presented to update the regional assignment of the gene for lactate dehydrogenase A (LDHA) to 11p12 leads to 11p13, and to support a previous assignment of the genes for the two components of the human cell-surface antigens of the SA11 (previously designated AL) group, SA11-1 and SA11-3 (previously designated AL-a1 and AL-a3), to 11pter leads to 11p13. This regional clone panel will be useful for rapid regional mapping of other genes assigned to chromosome 11. Show less

beta-Glucuronidase (GUS) has become an important enzyme model for the genetic study of molecular disease, enzyme realization, and therapy, and for the biogenesis and function of the lysosome and lysosomal enzymes. The genetics of human beta-glucuronidase was investigated utilizing 188 primary man-mouse and man-chinese hamster somatic cell hybrids segregating human chromosomes. Cell hybrids were derived from 16 different fusion experiments involving cells from ten different and unrelated individuals and six different rodent cell lines. The genetic relationship of GUS to 28 enzyme markers representing 19 linkage groups was determined, and chromosome studies on selected cell hybrids were performed. The evidence indicates that the beta-glucuronidase gene is assigned to chromosome 7 in man. Comparative linkage data in man and mouse indicate that the structural gene GUS is located in a region on chromosome 7 that has remained conserved during evolution. Involvement of other chromosomes whose genes may be important in the final expression of GUS was not observed. A tetrameric structure of human beta-glucuronidase was demonstrated by the formation of three heteropolymers migrating between the human and mouse molecular forms in chromosome 7 positive cell hybrids. Linkage of GUS to other lysosomal enzyme genes was investigated. beta-Hexosaminidase (HEXB) was assigned to chromosome 5; acid phosphatase2 (ACP2) and esterase A4 (ES-A4) were assigned to chromosome 11; HEXA was not linked to GUS; and alpha-galactosidase (alpha-GAL) was localized on the X chromosome. These assignments are consistent with previous reports. Evidence was not obtained for a cluster of lysosomal enzyme structural genes. In demonstrating that GUS was not assigned to chromosome 9 utilizing an X/9 translocation segregating in cell hybrids, the gene coding for human adenylate kinase1 was confirmed to be located on chromosome 9. Show less

A procedure is described which permits the isolation from the prepuberal mouse testis of highly purified populations of primitive type A spermatogonia, type A spermatogonia, type B spermatogonia, preleptotene primary spermatocytes, leptotene and zygotene primary spermatocytes, pachytene primary spermatocytes and Sertoli cells. The successful isolation of these prepuberal cell types was accomplished by: (a) defining distinctive morphological characteristics of the cells, (b) determining the temporal appearance of spermatogenic cells during prepuberal development, (c) isolating purified seminiferous cords, after dissociation of the testis with collagenase, (d) separating the trypsin-dispersed seminiferous cells by sedimentation velocity at unit gravity, and (e) assessing the identity and purity of the isolated cell types by microscopy. The seminiferous epithelium from day 6 animals contains only primitive type A spermatogonia and Sertoli cells. Type A and type B spermatogonia are present by day 8. At day 10, meiotic prophase is initiated, with the germ cells reaching the early and late pachytene stages by 14 and 18, respectively. Secondary spermatocytes and haploid spermatids appear throughout this developmental period. The purity and optimum day for the recovery of specific cell types are as follows: day 6, Sertoli cells (purity>99 percent) and primitive type A spermatogonia (90 percent); day 8, type A spermatogonia (91 percent) and type B spermatogonia (76 percent); day 18, preleptotene spermatocytes (93 percent), leptotene/zygotene spermatocytes (52 percent), and pachytene spermatocytes (89 percent), leptotene/zygotene spermatocytes (52 percent), and pachytene spermatocytes (89 percent). Show less

A reliable and uniform vascular perfusion fixation method for the testis has been developed by using an initial washout solution containing a vasodilator and an anticoagulant. This is followed by a brief fixation with a sodium phosphate buffered formaldehyde-glutaraldehyde solution of conventional strenght, and then a second more concentrated aldehyde fixative solution containing picric acid. The method takes into account some of the unique features of the vascular supply of the male genital tract for its favorable perfusion and fixation. The advantages of this method are: (1) consistently favorable preservation of the testis; (2) simple and inexpensive apparatus; and (3) stable and relatively innocuous stock solutions. Show less

Techniques of quantitative stereology have been utilized to determine the relative volume occupied by the Sertoli cells and germ cells in two particular stages (I and VII) of the cycle of the seminiferous epithelium. Sertoli cell volume ranged from 24% in stage I of the cycle to 32% in stage VII. Early germ cells occupied 3.4% in stage I (spermatogonia) and 8.7% in stage VII (spermatogonia and preleptotene spermatocytes). Pachytene spermatocytes occupied 15% (Stage I) and 24% (stage VII) of the total volume of the seminiferous epithelium. In stage I the two generations of spermatids comprised 58% of the total epithelium by volume, whereas in stage VII, after spermiation, the acrosome phase spermatids occupied 35% of the total seminiferous epithelial volume. Show less

The effect of caffeine on postreplication repair, as seen in alkaline sucrose gradients, conjugation, and ultraviolet light (UV) survival, was studied in excision deficient strains of Escherichia coli K12 and B. A caffeine concentration of 2 mg/ml was chosen for the study which did not inhibit colony formation. Both E. coli K12 AB2500 and E. coli B WWP2 were more sensitive to UV when plated on caffeine plates. Conjugation was not inhibited in the E. coli K12 strain; however, the same procedure confirmed caffeine inhibition in the E. coli B strain [17]. Caffeine did not inhibit postreplication repair in either strain, as determined by sedimentation profile studies of DNA on alkaline sucrose gradients. No strand breakage or degradation was observed in parental or post-UV replicated DNA for as long as 50 min incubation in caffeine. Thus caffeine concentrations that inhibited two recA gene product related phenomena did not cause immediate changes in size of DNA or inhibit the rate of a DNA gap generating postreplication type of DNA repair. Show less

The fine structure of the rete testis was examined in several primates, domestic animals and rodents. The rete testis consists of a series of interconnected wide channels lined with a simple cuboidal to columnar epithelium, resting on a thick basal lamina. Beneath the basal lamina dense bundles of collagen fibrils and a few blood vessels, lymphatics or nerve tissue are found. The epithelial cells are characterized by large, deeply indented nuclei, spherical or short rod-shaped mitochondria, supranuclear Golgi profiles, some cisterns of rough endoplasmic reticulum, free ribosomes and numerous micropinocytotic vesicles in the ectoplasmic regions. Smooth endoplasmic reticulum, secretory granules, lysosomes or other types of dense bodies are rarely seen. The apical surface of the cells bears numerous microvilli and a single very long flagellum which is presumed to be motile. Ajoining lateral cell membranes exhibit a juxtaluminal tight junction, elaborate interdigitations and desmosomes. The basal plasma membrane is highly irregular greatly increasing its surface area of contact with the underlying interstitium. The nuclei of the rete epithelial cells contain pale-staining, spherical structure, 2 mum in diameter, composed of circularly oriented fine filaments. The significance of the nuclear structures remains unknown. Thorotrast was injected into the lumen of the hamster and rat rete testis and 30 minutes later the proximal portion of the excurrent duct system of the testis was prepared for electron microscopy. Whereas the ductuli efferentes and first part of the epididymis possessed numerous apical vesicles filled with the thorotrast, this electron opaque substance was rarely found in the epithelium of the rete testis. Thus, incorporation of particulate matter into the lining cells of the rete from its lumen is apparently less active than in the epithelium of the ductuli and epididymis. Vascularly introduced intercellular tracer compounds such as lanthanum nitrate or horseradish peroxidase did not enter the lumen of the rete testis from the interstitium. The tracer molecules appeared to be blocked by the juxtaluminal tight junction separating adjacent epithelial cells. This latter observation suggests that a blood-testis barrier exists at the level of the rete testis epithelium. Although physiological studies have indicated that the composition of fluid secreted in the seminiferous epithelium is considerably modified in the rete testis, the present morphological study does not provide additional evidence to support a secretory or absorptive function for this region of the excurrent duct system of the testis. Show less

Epidemiological studies and clinical observation suggesting potential hazards of arsenic compounds in increasing the incidence of cancer have been in complete contradiction with experimental findings in animals. Because of the predominance of skin cancers in the epidemiological reports, we decided to investigate the possibility that arsenic compounds might interfere with DNA repair. Using Escherichia coli as a test system, we show that this is indeed the case. Sodium arsenite, at concentrations of 0.1 mM and higher, decreases the survival of ultraviolet-irradiated E. coli WP2, a strain which possesses the full complement of repair genes. The effect of the arsenite increases with increasing ultraviolet dose. Similar results were obtained with the excision repair deficient strains WWP2 (uvrA) and WP6 (polA). Sodium arsenite had no effect on the survival of a recA mutant, WP10. Survival of ultraviolet-irradiated WP5 (exrA) was enhanced by sodium ardenite, the effect being greatest at low ultraviolet doses. It is postulated that arsenite inhibits a recA-dependent step in DNA repair. To account for the increased survival of the exrA mutant, we suggest that in the absence of the exr+ gene, the arsenite-sensitive recA-dependent function is deleterious. The ability of arsenite to inhibit DNA repair may account for the clinical and epidemiological reports linking arsenicals with an increased incidence of cancer. Show less

When the excision proficient strain E. coli WP2 Hcr+ trp- was grown to stationary phase by glucose starvation in M-9 minimal medium before UV -irradiation, the ability of nutrient broth enrichment of minimal medium to enhance trp- leads to Trp+ reversion was greatly reduced. Less than 50% of the Trp+ revertants were found to be ochre suppressors. However, in the WWP2 Hcr- strain, 75-86% of the tested revertants were ochre suppressors. This indicates that, under the cultural conditions employed, many potential suppressor mutations were removed by excision repair in the presence of broth enrichment. Broth enhancement of reversion also occurred in the Hcr- strain, which indicates that a less error-prone mode of recombination repair functions under minimal growth conditions. An Hcr+ strr derivative of WP2 Hcr+ was more resistant than its strs parent to the lethal effect of UV light and showed a lower UV-induced Trp+ reversion frequency. The percentage of Trp+ revertants that were due to ochre suppressors was markedly reduced in the strr strain. The Hcr- strr strain also had a lower UV-induced Trp+ reversion frequency than its strs parent. The excision repair inhibitor caffeine had little effect at lower UV doses on increasing Trp+ reversion in both Hcr+ strains. Acriflavine, however, was effective at lower UV doses in enhancing reversiin of the Hcr+ strains and the degree of enhancement increased with the dose. Acriflavine appeared to specifically enhance the number of ochre suppressing Trp+ revertants. In both Hcr- strains (strs and strr) caffeine (500 mug/ml) had no effect on survival but reduced the UV-induced Trp+ reversion frequency acting as an antimutagen. In contrast, acriflavine (2 mug/ml) decreased survival and increased the Trp+ reversion frequency of the Hcr- strains. The data on spontaneous Trp+ reversion frequencies show that the Hcr+ strs strain had a higher spontaneous reversion frequency than the Hcr- strs strain on all plating media. Further, caffeine was shown to reduce spontaneous Trp+ reversion in both Hcr+ and Hcr- strains while acriflavine increased the spontaneous reversion frequencies of both strains. Show less