Side population cells in human and mouse epidermis lack stem cell characteristics

Cells that exclude Hoechst 33342 have been found in many tissues, and common for these cells is a characteristic profile when analysed by flow fluorimetry (sp, side population). Since sp cells in some cases function as multipotent stem cells, we investigated whether the epidermis contains sp cells (Esp cells) and whether these cells were epidermal stem cells. We show that mouse and human epidermis contain sp cells, and, to identify the origin of these cells, we tested the expression of several marker genes. We find that Esp cells constitute a subpopulation of the alpha6 integrin-positive basal cells of the mouse epidermis. They are positive for sca-1 and negative for MHC class II and Flk1. They are not identical to the label-retaining population but are cycling cells in the mouse epidermis. Keratinocytes positive for sca-1 are located outside the stem cell containing bulge area of the mouse hair follicle. Forty-four human skin samples were analysed, and Esp cells were found at frequencies ranging from 0.01% to 5.39%, independently of age and body site. Human Esp cells did not express particular high levels of beta1 integrin. However, they expressed the half transporter ABCG2 and we identified high expression of this marker in the secretory duct epithelium of the sweat glands whereas low expression was found in the basal layer of the epidermis.     

Gramicidin-perforated patch recording revealed the oscillatory nature of secretory Cl- movements in salivary acinar cells

Elevations of cytoplasmic free calcium concentrations ([Ca(2+)](i)) evoked by cholinergic agonists stimulate isotonic fluid secretion in salivary acinar cells. This process is driven by the apical exit of Cl(-) through Ca(2+)-activated Cl(-) channels, while Cl(-) enters the cytoplasm against its electrochemical gradient via a loop diuretic-sensitive Na(+)-K(+)-2Cl(-) cotransporter (NKCC) and/or parallel operations of Cl(-)-HCO(3)(-) and Na(+)-H(+) exchangers, located in the basolateral membrane. To characterize the contributions of those activities to net Cl(-) secretion, we analyzed carbachol (CCh)-activated Cl(-) currents in submandibular acinar cells using the "gramicidin-perforated patch recording configuration." Since the linear polypeptide antibiotic gramicidin creates monovalent cation-selective pores, CCh-activated Cl(-) currents in the gramicidin-perforated patch recording were carried by Cl(-) efflux via Cl(-) channels, dependent upon Cl(-) entry through Cl(-) transporters expressed in the acinar cells. CCh-evoked oscillatory Cl(-) currents were associated with oscillations of membrane potential. Bumetanide, a loop diuretic, decreased the CCh-activated Cl(-) currents and hyperpolarized the membrane potential. In contrast, neither methazolamide, a carbonic anhydrase inhibitor, nor elimination of external HCO(3)(-) had significant effects, suggesting that the cotransporter rather than parallel operations of Cl(-)-HCO(3)(-) and Na(+)-H(+) exchangers is the primary Cl(-) uptake pathway. Pharmacological manipulation of the activities of the Ca(2+)-activated Cl(-) channel and the NKCC revealed that the NKCC plays a substantial role in determining the amplitude of oscillatory Cl(-) currents, while adjusting to the rate imposed by the Ca(2+)-activated Cl(-) channel, in the gramicidin-perforated patch configuration. By concerting with and being controlled by the cation steps, the oscillatory form of secretory Cl(-) movements may effectively provide a driving force for fluid secretion in intact acinar cells.     

Wing morph-related differences in developmental pattern of accessory gland proteins in adult males of Pyrrhocoris apterus (L.) and their endocrine control

The study showed that the amounts of the total proteins and 53 kDa protein in male accessory glands (AGs) of the firebug Pyrrhocoris apterus (L.) increased with age of the adult life. The 53 kDa protein, the most abundant polypeptide detected in the secretion of the AGs, and some other smaller peptides were identified as glycoproteins. Changes in the amounts of the total proteins and 53 kDa protein in AGs were found to be wing morph-dependent and their levels were significantly higher in 1-10 days old brachypterous males than in macropterous males of the same age. Macropterous males were characterized by delayed growth of the AGs. Treatment of adult macropterous males with methoprene significantly increased the amounts of total proteins and 53 kDa protein in their AGs when compared to acetone-treated macropterous controls of the same age. Allatectomy of brachypterous males decreased the levels of the total proteins and 53 kDa protein in their AGs, while application of methoprene enhanced the quantity of allatectomy-suppressed proteins in these tissues. This is the first report of juvenile hormone-dependent wing morph-related differences in the synthesis of AG proteins and their endocrine control in wing-polymorphic insects.     

Vasculature of the parotoid glands of four species of toads (bufonidae: bufo)

The parotoid glands of toads (Bufonidae) consist of large aggregations of granular glands located between the otic region of the skull and the scapular region. To determine the circulatory pattern of these glands, we perfused the vascular systems of Bufo alvarius, B. marinus, B. terrestris, and B. valliceps with either India ink or Microfil, a fine latex. The perfused glands were studied by gross dissection, microscopic examination, and histology. The vascular patterns of the parotoid glands were compared to the arrangement of vessels in the dorsal skin of Rana sphenocephala (Ranidae), a frog that lacks parotoid glands. The parotoid glands of the four species of toads are supplied with blood by the lateral and dorsal cutaneous arteries and are drained by one or more branches of the internal jugular vein. The dorsal cutaneous artery supplies most of the blood to the parotoid glands in B. terrestris and B. valliceps. In B. alvarius and B. marinus, both the lateral and dorsal cutaneous arteries serve major roles in the blood supply of the glands. These patterns of blood flow have not been described previously for parotoid glands and conflict with earlier accounts for B. alvarius and B. marinus. The arteries and veins associated with the parotoid glands of toads are present in R. sphenocephala, but are arranged differently. In R. sphenocephala, the lateral cutaneous artery supplies the dorsal and lateral skin posterior to the shoulder region, whereas the dorsal cutaneous artery supplies the skin of the shoulder region. In toads, both the lateral and dorsal cutaneous arteries supply the skin of the shoulder region and ramify into subcutaneous capillaries that surround the secretory units of the parotoid glands. Extensive vasculature presumably is important for delivering cholesterol and other precursor molecules to the parotoid glands, where those compounds are converted into toxins.     

Ultrastructure of posterior sternal glands of Macrotermes annandalei (Silvestri): new members of the sexual glandular set found in termites (Insecta)

In female alates of Macrotermes annandalei, two types of abdominal glands are involved in the secretion of sex pheromone. Tergal glands are found at the anterior margin of tergites 6-10 and posterior sternal glands (PSGs) are located at the anterior margin of sternites 6-7. The cytological features of both types of glands are quite similar. The fine structural organization of PSGs is studied more precisely and described for the first time. The glandular cuticle is pitted with narrow apertures corresponding to the openings of numerous subcuticular pouches. Several Class 3 glandular units open in each pouch. One canal cell and one secretory cell make an individual glandular unit. The canal cell is enlarged apically and is connected with the other canal cells to form a common pouch. Based on the structural features found in these glands, we propose a common secretory process for PSGs and tergal glands. During the physiological maturation of alates inside the nest, secretory vesicles amass in the cytoplasm of secretory cells, while large intercellular spaces collapse the cuticular pouches. At the time of dispersal flight, pouches are filled with the content of secretory vesicles while intercellular spaces are sharply reduced. After calling behavior, no secretion remains in the glands and pouches collapse again, while secretory cells are drastically reduced in size. The structure and the secretory processes of PSGs and tergal glands are compared to those of abdominal sexual glands known in termites.     

Evolutionary transformation from muscular to hydraulic movements in spider (Arachnida, Araneae) genitalia: a study based on histological serial sections

The male genitalia of 107 spider species representing 73 families were serially sectioned and studied with an emphasis on muscles moving the genital bulb. As a rule, most non-Entelegynae have two bulbal muscles, most Entelegyne have none, but many exceptions occur. Variation also occurs with regard to origin and attachment of bulbal muscles. There appears to be a trend towards a shift of the origin from proximal (Liphistius, Atypus) to more distal palpal segments (Haplogynae). In most Entelegynae the muscular movement is replaced by hydraulic movement caused by expanding membranes (hematodochae). Hematodochae probably permit increased bulbal rotation and movements of higher complexity. New evidence is presented arguing against Palpimanidae being representatives of Entelegynae. Bulbal glands other than those discharging into the sperm duct (previously known in Amaurobiidae and Dictynidae only) are described in several entelegyne families.     

Molecular evolution and population genetics of duplicated accessory gland protein genes in Drosophila

To investigate the potential importance of gene duplication in D. melanogaster accessory gland protein (Acp) gene evolution we carried out a computational analysis comparing annotated D. melanogaster Acp genes to the entire D. melanogaster genome. We found that two known Acp genes are actually members of small multigene families. Polymorphism and divergence data from these duplicated genes suggest that in at least four cases, protein divergence between D. melanogaster and D. simulans is a result of directional selection. One putative Acp revealed by our computational analysis shows evidence of a recent selective sweep in a non-African population (but not in an African population). These data support the idea that selection on reproduction-related genes may drive divergence of populations within species, and strengthen the conclusion that Acps may often be under directional selection in Drosophila.     

Enigmas of mammalian gamete form and function

The gametes of man and some other Eutheria have been manipulated successfully for practical reasons, but many gaps remain in our basic understanding of the way that they function. This situation stems not least from a failure to recognize the extent to which eutherian spermatozoa and eggs, and elements related to their operation, have come to differ from those of other groups. Novel features in the male that reflect this include a radical design of the sperm head with the acrosome seeming to function primarily in egg-coat binding rather than its lysis, a multifaceted post-testicular sperm maturation and an androgen/low-temperature-regulated system of sperm storage--both tied to the epididymis, a variable male accessory sex gland complex, and descent of the testis and epididymis to a scrotum. In the female, such novelties are represented in a need for sperm capacitation, in an unusual regulation of sperm transport within the oviduct, in the cumulus oophorus and character of the zona pellucida around the small egg, and in a unique configuration of gamete fusion. The collective evidence now suggests that many of these features reflect a new fertilisation strategy or its consequences, with most being causally linked. One initial 'domino' in this regard appears to be the small yolkless state of the egg and its intolerance for polyspermy, as determinants of the unusual mode of oviductal sperm transport and possibly the existence and form of the cumulus oophorus. However, a particularly influential first 'domino' appears to be the physical character of the eutherian zona pellucida. This differs from the egg coats of other animal groups by virtue of a resilient elasticity and thickness. These qualities allow this primary and often only coat to stretch and so persist during later expansion of the blastocyst, usually until close to implantation. At the same time, the dimensions, physical character, and particularly the relative protease-insensitivity of the zona appear to have had profound effects on sperm form and function and, more indirectly, on sperm-related events in the male and the female tract. Marsupials display some similarities and also some strikingly different features, against which the enigmas of the eutherian situation can be evaluated.