Quantification of human spermatogenesis: germ cell degeneration during spermatocytogenesis and meiosis in testes from younger and older adult men

Germ cell degeneration during spermatocytogenesis and meiosis was investigated to explain the age-related decline in daily sperm production (DSP). Numbers of Types A-dark, A-pale, and B-spermatogonia, potential daily sperm production per g parenchyma (PDSP) based on type B-spermatogonia, early primary spermatocytes, and late primary spermatocytes, and DSP per g based on early spermatids were determined in 15 men aged 20 to 48 yr (mean +/- SEM, 33 +/- 2 yr) and 15 men aged 52 to 90 yr (65 +/- 3 yr). Testes obtained within 15 h of death (largely due to trauma or heart failure) were perfused vascularly with glutaraldehyde. The number of each cell type per g parenchyma was calculated as the product of the percentage of nuclei in the parenchyma times a correction factor for section thickness and nuclear diameter divided by the volume of a single nucleus of that cell type. Paired testicular weight was lower (p less than 0.01) in older men (33 +/- 3 g) than in the younger men (49 +/- 3 g). Younger and older men had similar numbers of A-dark, A-pale, and B-spermatogonia per g parenchyma. PDSP based on late primary spermatocytes and DSP based on early spermatids were lower (p less than 0.01) in older men than in younger men. In younger men, PDSP was similar (p greater than 0.05) between B-spermatogonia and late primary spermatocytes, whereas DSP measured at the spermatid level was abruptly lower than that estimated from younger cell types. Older men showed reduction in PDSP between early and late primary spermatocytes, with further reduction occurring in DSP at the spermatid level.(ABSTRACT TRUNCATED AT 250 WORDS)     

Trichomoniasis in free-living goshawks (Accipiter gentilis gentilis) from Great Britain

The goshawk Accipiter gentilis has recently been reintroduced into parts of Great Britain. During the course of a study of one population, lesions of stomatitis were observed in 14 young from five broods and all the affected birds died. Postmortem examination of three birds revealed live Trichomonas gallinae in exudate from one, and histological findings consistent with a diagnosis of trichomoniasis were made in this and one other bird. It is suggested that trichomoniasis may be a significant mortality factor in goshawks from Britain.     

Do male hoots betray parasite loads in Tawny Owls?

Bird song structure may honestly reveal the health and vigour of individual males to potential mates and competitors. If this is the case then song may reflect the level of parasitic infections in males. We initially examined the relationship between blood parasite infections and the time taken to respond by 22 male Tawny Owls to a broadcast hoot. We then examined the call structure (total length and frequency) in relation to parasite infection, an index of owl condition and an index of food abundance. Owls with higher parasite loads responded more slowly to an intruder, although this relationship was not significant once condition and vole abundance were controlled for. We found no relationship between call length and any of the measured variables. However, the high frequency and the range of frequencies used in calls decreased with increasing parasite load. Thus, there was the potential for individuals to assess male parasite load from the speed of response and the structure of the call. Experimental tests of these relationships are now required.     

Influence of immune complexes on macrophage membrane fluidity: a nanosecond fluorescence anisotropy study

Time-resolved fluorescence anisotropy (TRFA) and steady-state anisotropy measurements and fluorescence intensification microscopic observations were made on RAW264 macrophages labeled with 1,6-diphenyl-1,3,5-hexatriene (DPH) or 1-[4-(trimethylammonio)phenyl]-6-phenyl-1,3,5-hexatriene (TMA-DPH). Microscopic analysis revealed that the fluorescent probe DPH was found in association with plasma membranes and small vesicles. Macrophages treated with immune complexes could not be distinguished from untreated cells, indicating that the same membrane compartments were labeled. The probe TMA-DPH was exclusively localized to the plasma membrane. Steady-state anisotropy measurements indicated that in vitro culture conditions did not significantly affect membrane fluidity. TRFA measurements were conducted to determine the physical properties of macrophage membranes during immune recognition and endocytosis. Data were analyzed by iterative deconvolution to yield phi, the rotational correlation time, and r infinity, the limiting anisotropy. These parameters may be interpreted as the "fluidity" and order parameter of the membrane environment, respectively. Typical values for untreated macrophages were phi = 7.8 ns and r infinity = 0.12. Binding and endocytosis of immune complexes prepared in 4-fold antigen excess increase these values to phi = 22.1 ns and r infinity = 0.15. However, receptor-independent phagocytosis of latex beads decreases these values to phi = 2.2 ns and r infinity = 0.10. Addition of catalase before, but not after, immune complex incubation with cells diminishes the effect upon membrane structure, suggesting that H2O2 participates in fluidity changes. Pretreatment of macrophages with the membrane-impermeable sulfhydryl blocker p-(chloromercuri)benzenesulfonic acid also diminished these effects.(ABSTRACT TRUNCATED AT 250 WORDS)     

Micro-scission of YAC tumor cells during antibody-dependent cellular cytotoxicity mediated by human neutrophils

The cellular events accompanying neutrophil-mediated antibody-dependent cellular cytotoxicity (ADCC) directed against YAC erythroleukemic target cells have been studied by time-lapse fluorescence-intensified microscopy. The YAC plasma membrane and cytosol were labeled with the fluorescent probes diC18Icc and eosin Y, respectively. Fluorescently labeled and IgG-opsonized YAC cells were incubated at 37 degrees C while observed by optical microscopy. During temporal studies of neutrophil-YAC conjugates, the cytosol of YAC cells accumulated in tubular and spherical compartments of the neutrophils' vacuolar apparatuses. To distinguish between several possible mechanisms of target cytosol uptake, diC18Icc-labeled YAC cells were observed during identical conditions. The membrane label diC18Icc was found to accumulate within neutrophils in an identical fashion. At roughly 30 min, 25 and 38% of neutrophils in apparent conjugates had internalized tumor cell cytosol or plasma membrane, respectively, within a vesicular compartment. The IgG-dependent uptake of eosin Y and diC18Icc by neutrophils was diminished by exposure to 2.5 mM sodium azide. When cells were exposed to 5.5 mM sodium azide, 1 mM iodoacetamide, or 4 degrees C, conjugate formation and uptake of eosin Y or diC18Icc were abolished. An artifactual accumulation of eosin Y or diC18Icc in neutrophils was further ruled out by control studies. Non-specific exchanges of eosin Y and diC18Icc labels of YAC cells with tannic acid-treated red blood cells (RBCs) and normal neutrophils were studied. Since hemoglobin binds tightly to eosin Y, RBCs can easily detect eosin Y leakage. No exchange of eosin Y or diC18Icc from YAC cells into bound tannic acid-treated erythrocytes was found.(ABSTRACT TRUNCATED AT 250 WORDS)     

Behavioral responses of Diabrotica adults to plant-derived semiochemicals encapsulated in a starch borate matrix

The concept of encapsulating semiochemicals into a starch matrix is being studied for potential use in corn rootworm (CRW) management programs. During 1987, experiments were conducted to determine: 1) If volatile plant-derived Diabrotica spp. attractants could be encapsulated in a starch borate matrix (SBM), and 2) If various SBM-semiochemical formulations would attract Diabrotica species over time in field corn. Chemical analyses of fresh SBM formulations indicated that indole, estragole, veratrole, phenylacetaldehyde, and trans-anethole were not retained during formulation but trans-cinnamaldehyde, Beta-ionone, 1,2,4,-trimethoxybenzene, eugenol and isugenol were successfully encapsulated. Encapsulated semiochemical formulations were made into 20 mesh granules, placed in Pherocon ® 1C traps that were tied to corn plants, and sampled for CRW adults every 4 days from 11 July to 8 September. Field data indicated that encapsulated semiochemicals were retained in the SBM for varying lengths of time and were released at rates attractive to CRW adults. A two-component mixture of trans-cinnamaldehyde and 1,2,4-trimethoxybenzene was the most effective formulation tested; however, no formulation was effective during corn silking and pollination. Although seasonal variation in CRW response could limit the usefulness of some plant-derived semiochemicals, the starch matrix concept may be useful as a delivery system for semiochemicals and may have potential as a tool that could be used in the development of new more biorational CRW management programs.

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Résumé Le programme expérimental de 1987 était destiné à déterminer: 1) si les substances dérivées de végétaux et attractives pour Diabrotica pouvaient être encapsulées dans de l'amidon additionné d'acide borique; 2) si différentes formules attireraient les différentes espèces de Diabrotica dans un champ de maïs.L'indol, l'estragol, le vératrol, le phénylacétaldéhyde et le trans-anéthol n'ont pas été retenus, tandis que le trans-cinnamaldéhyde, la \-ionone, le 1,2,4-triméthobenzène, l'eugénol et l'isugénol ont été encapsulés avec succès dans des pièges attachés à des pieds de maïs (les détails techniques sont fournis). Les pièges ont été relevés tous les 4 jours du 11 juillet au 8 septembre. Les résultats montrent que les substances allélochimiques sont conservées dans la capsule pendant des durées variables et libérées à des concentrations attractives pour les Diabrotica adultes. Un mélange de trans-cinnamaldéhyde et de 1,2,4,-triméthoxybenzène a été la formule la plus efficace, à l'exception des périodes de formation des barbes et du pollen, où aucune formule n'a été attractive. Bien que la variation saisonnière des réactions de Diabrotica limite l'utilisation des substances allélochimiques d'origine végétale, la capsule d'amidon peut être employée pour libérer des substances allélochimiques et constitue un outil potentiel pour la mise au point d'une méthode plus rationnelle de lutte contre Diabrotica.

     

Isolation and regional localization of a large collection (2,000) of single-copy DNA fragments on human chromosome 3 for mapping and cloning tumor suppressor genes

Summary A collection of 2,000 lambda phage-carrying human single-copy inserts (> 700 bp) were isolated from two chromosome-3 flow-sorted libraries. The single-copy DNA fragments were first sorted into 3p and 3q locations and about 700 3p fragments were regionally mapped using a deletion mapping panel comprised of two humanhamster and two-human-mouse cell hybrids, each containing a chromosome 3 with different deletions in the short arm. The hybrids were extensively mapped with a set of standard 3p markers physically localized or ordered by linkage. The deletion mapping panel divided the short arm into five distinct subregions (A-E). The 3p fragments were distributed on 3p regions as follows: region A, 26%; B, 31%; C, 4%; D, 4% and E, 35%. We screened 300 single-copy DNA fragments from the distal part of 3p (regions A and B) with ten restriction endonucleases for their ability to detect restriction fragment length polymorphisms (RFLPs). Of these fragments 110 (36%) were found to detect useful RFLPs: 35% detected polymorphisms with frequency of heterozygosity of 40% or higher, and 25% with frequency of 30% or higher. All polymorphisms originated from single loci and most of them were of the base pair substitution type. These RFLP markers make it possible to construct a fine linkage map that will span the distal part of chromosome 3p and encompasses the von Hippel-Lindau disease locus. The large number of single-copy fragments (2,000) spaced every 100–150 kb on chromosome 3 will make a significant contribution to mapping and sequencing the entire chromosome 3. The 300 conserved chromosome 3 probes will increase the existing knowledge of man-mouse homologies.     

Cytochemical study of macrophage lysosomal inorganic trimetaphosphatase and acid phosphatase

Cytochemical investigations have associated acid inorganic trimetaphosphatase (TMPase) activity with the lysosomes of certain cell types. We have used the modified staining technique of Berg to show that this enzyme activity is present in normal mononuclear phagocytes and macrophage cell lines. We have found this enzyme activity to be present in murine RAW264 macrophages, in human U937 macrophages, in normal human blood monocytes, and in guinea pig peritoneal macrophages. All of the RAW264 and U937 macrophages showed intense TMPase activity. Many of the human monocytes and most of the guinea pig macrophages were labeled by this method. The reaction product was associated with the lysosomes of these cell types. The lysosomal staining-pattern was similar to that of acid phosphatase. Differences with regard to Golgi staining were noted. This indicates that TMPase is a lysosomal enzyme of mammalian macrophages. The distinction between TMPase and acid phosphatase activity has been demonstrated by measuring the pH optimum of each enzyme. Using substrates identical to those of the ultrastructural cytochemistry, we show that the pH optimum of TMPase is 4.0 and that of acid phosphatase is 5.0. The enzymatic activities are therefore ultrastructurally and biochemically distinct. Following phagocytosis of latex, yeast (Saccharomyces cerevisiae), or Corynebacterium parvum, TMPase has been found to be associated with phagosomes. This enzyme may take part in the degradation of phagocytosed materials, particularly microorganisms which contain inorganic polyphosphates and metaphosphates.     

Influence of age on sperm production and testicular weights in men

Age-related changes in daily sperm production (DSP) and testicular weights were investigated in paired testes from 89 men aged 21-50 years and 43 men aged 51-80 years. For both DSP/testis and DSP/g parenchyma, remarkably large standard deviations exceeded 50% of mean values. However, DSP/g and DSP/testis for both right and left testes were approximately 30% higher in the younger than in the older group (P less than 0.01) and were negatively correlated with age (P less than 0.01) when data from both groups were pooled. Weights of whole testes and of testicular parenchyma were similar in both age groups and were not significantly correlated with age. However, testicular tunic weights were 29% higher in the older group (P less than 0.001) and were positively correlated with age (P less than 0.001). Both testicular tunic weight and the % of total testis occupied by tunic were negatively correlated with DSP/g (P less than 0.01); these correlations were weakened by removing the effect of age. Although total testicular weight and testicular parenchymal weight did not change with age, these values were about 10% lower on the left than on the right (P less than 0.001). In addition to its increase with age, testicular tunic weight was about 8% greater for right than for left testes in all men (P less than 0.001). Although the average size of the testis varied from right to left, DSP/g was similar in paired testes (P = 0.15), and the correlation between right and left DSP/g was high (rho = +0.89, P less than 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)