The formation of intercellular junctions in insect stem cell progeny (cockroach intestinal epithelium)

Summary The stages in the development of intercellular junctions have been followed in the mesenteric caecal cells of the cockroach midgut, where two types of mature cell, the columnar and the secretory, exist. Nests of undifferentiated replacement cells occur at intervals along the basal lamina, consisting of central, dividing cells and peripheral semi-lunar cells; the former act as proliferative stem cells to give rise to either pre-columnar or pre-secretory cells. The semi-lunar cells are pre-columnar and produce an attenuated process which gradually projects up to the luminal surface, producing microvilli and a dense extracellular substance en route. Intercellular gap junctions appear between these maturing columnar cell borders first, while septate junctions differentiate later; these are assembled from two different sets of intramembranous particle which become organized into either plaques or rows in parallel alignment, possibly mediated by actin filaments and microtubules. The pre-secretory cells, which are much fewer in number, remain associated only with the basal lamina and never reach the lumen; they develop into one of three distinct mature secretory cell types which release their secretory product in different ways. Offprint requests to: N.J. Lane     

线纹香茶菜叶上腺毛发育的细胞学研究

为进行中药溪黄草基原植物的品种鉴定,采用光镜和电镜对线纹香茶菜(原变种)[Isodon lophanthoides var.lophanthoides]叶上腺毛的发育进行细胞学研究。结果表明,线纹香茶菜具有头状腺毛和盾状腺毛2种类型。头状腺毛无色透明,由1个基细胞、1个柄细胞和1或2个头部分泌细胞构成;盾状腺毛为红色,由1或2个基细胞、1个柄细胞和4~8个分泌细胞构成头部。2种腺毛均由原表皮细胞经两次平周分裂形成,后因柄细胞和头部细胞所处的分化状态不同而形成两类腺毛。2种腺毛超微结构表明,质体、高尔基体和粗面内质网为主要分泌物产生和运输的细胞器。当盾状腺毛成熟时,角质层下间隙充满了分泌物,其分泌物的性质很可能决定了线纹香茶菜腺毛的颜色。     

Accumulation and secretion of exoglucanase activity in yeast secretory mutants

Representative conditional yeast secretory mutants, blocked in transport of secretory and plasma membrane proteins from the endoplasmic reticulum (sec 18), from the Golgi body (sec 7) and in transport of secretory vesicles (sec 1), accumulated exoglucanase, a constitutive yeast activity, when incubated at the restrictive temperature (37°C). Different proportions of the accumulated activity were released by mutant cells under permissive conditions. The presence or absence of cycloheximide during the secretion period made no differences in the results. More than 90% of the internal activity was bound to membrane in wild type cells. However, only the soluble pool underwent changes during the accumulation or secretion periods. The bulk of secretory invertase accumulated by sec 1 was also soluble. By contrast sec 7 and sec 18 accumulated membrane-bound as well as soluble invertase forms and both were secreted in similar proportions in each mutant. More than 90% of the accumulated invertase was secreted at the permissive temperature in sec 18 cells. That percentage was significantly lower for exoglucanase (<65%). Concomitantly, invertase accumulated by this mutant exited from the cells with a lower half time (t 1/2=150 min). These results may be interpreted assuming that exoglucanase is exported by a passive flow of the soluble pool.Non-standard abbreviations p-NPG p-nitrophenyl--d-glucopyranoside - Con A concanavalin A - Tris tris(hydroxymethyl)-amino-methane     

杭白芷根中分泌道发生方式、分布及其挥发油积累过程研究

为探明杭白芷(Angelica dahurica var.formosana)根中分泌道发生方式、分布及其挥发油转运积累特征,该研究利用光镜及透射电子显微镜技术观察分泌道发生过程及挥发油转运特征,结合组织化学定位确定挥发油的主要积累部位。结果表明:杭白芷根中分泌道由中柱鞘细胞最先发生,次生结构中分泌道主要分布在韧皮部和皮层中;挥发油的合成不仅与分泌细胞中质体及细胞质有关,而且还与周围细胞关系密切;分泌细胞内高尔基体和内质网丰富,可能先通过形成小泡参与转运,再经由细胞壁向腔道内转移;相邻分泌细胞靠近角隅处的细胞壁分泌活动活跃,腔道内积累大量电子致密物质;成熟分泌道中分泌细胞及其腔道内积累大量油滴,因此挥发油主要积累场所为分泌细胞及其腔道。该研究明确了杭白芷根中分泌道的发生方式、分布及其挥发油积累部位,揭示了分泌道发育过程中挥发油的转运积累特征,为进一步阐明分泌组织生长发育与有效成分积累关系提供了理论依据。     

龙葵腺毛发育的细胞学研究

利用光学显微镜、扫描电镜和透射电镜技术,观察了龙葵“四叶一心”期时叶片及茎表皮的腺毛的种类、分布,探究了不同类型腺毛的起源、生长、成熟、分泌、衰老等发育过程的细胞学特征;通过组织化学染色和荧光显微技术,观察了龙葵腺毛成分、分布,为龙葵的进一步开发利用提供参考。结果表明：（1）龙葵腺毛分为单细胞头腺毛和多细胞头腺毛两类,前者主要分布于茎表面和叶上下表皮,后者主要分布于茎表面的单细胞头腺毛之间、叶脉及叶边缘;（2）龙葵腺毛发育起始于表皮细胞突起,单细胞头腺毛行顶端生长,具1-4个柄细胞,四种类型;多细胞头腺毛可再分为一层、两层与三层多细胞头腺毛,另具三种特殊类型;（3）龙葵成熟腺毛具分泌能力,通过皮下空间的物质积累导致腺毛头细胞表面形成突起、包块、破口,最终释放分泌物;而头细胞与柄细胞随即皱缩、衰老。（4）超微结构显示,腺毛头细胞中内质网与高尔基体极为丰富,合成代谢及分泌活动活跃,产生大量包裹嗜锇物质的囊泡,囊泡与细胞壁融合,进而将嗜锇物质转移至细胞壁并积累,随后储存在角质层下的皮下空间直至分泌释放;（5）组织化学染色结果表明,腺毛含有萜类、生物碱、脂类、蛋白质、酚类和多糖。头细胞中主要含有萜类、生物碱、脂类、蛋白质、酚类和中性多糖;柄细胞中主要含有萜类、生物碱、脂类。     

贯叶连翘的分泌结构及其与金丝桃素积累的关系

贯叶连翘（ＨｙｐｅｒｉｃｕｍｐｅｒｆｏｒａｔｕｍＬ．）地上器官分布着分泌细胞球（黑色腺点）、分泌囊（半透明腺点）和分泌道（半透明腺条）３类内部分泌结构。分泌细胞球在茎、叶和花器官中均有分布,由２层鞘细胞包围多个分泌细胞构成实心的分泌细胞团。分泌囊主要分布于叶片中,分泌道则分布于花器官中,它们都是由１～２层切向扁平细胞围绕圆形或长形腔道构成,腔道的贮存物为精油。利用组织化学方法,结合荧光显微镜观察,证实金丝桃素类物质是由分泌细胞球（黑色腺点）所合成和积累的。通过用戊二醛和锇酸固定样品的显微和超微结构观察,发现金丝桃素类物质积累在成熟腺体分泌细胞的中央大液泡中,细胞周围浓厚的细胞质中分布着大量小液泡和高尔基体、内质网等细胞器。在此基础上对金丝桃素类物质的积累过程进行了初步探讨     

Morphology and ultrastructure of the venom glands of the northern pacific rattlesnake Crotalus viridis oreganus

The venom gland of Crotalus viridis oreganus is composed of two discrete secretory regions: a small anterior portion, the accessory gland, and a much larger main gland. These two glands are joined by a short primary duct consisting of simple columnar secretory cells and basal horizontal cells. The main gland has at least four morphologically distinct cell types: secretory cells, the dominant cell of the gland, mitochondria-rich cells, horizontal cells, and “dark” cells. Scanning electron microscopy shows that the mitochondria-rich cells are recessed into pits of varying depth; these cells do not secrete. Horizontal cells may serve as secretory stem cells, and “dark” cells may be myoepithelial cells. The accessory gland contains at least six distinct cell types: mucosecretory cells with large mucous granules, mitochondria-rich cells with apical vesicles, mitochondria-rich cells with electron-dense secretory granules, mitochondria-rich cells with numerous cilia, horizontal cells, and “dark” cells. Mitochondria-rich cells with apical vesicles or cilia cover much of the apical surface of mucosecretory cells and these three cell types are found in the anterior distal tubules of the accessory gland. The posterior regions of the accessory gland lack mucosecretory cells and do not appear to secrete. Ciliated cells have not been noted previously in snake venom glands. Release of secretory products (venom) into the lumen of the main gland is by exocytosis of granules and by release of intact membrane-bound vesicles. Following venom extraction, main gland secretory and mitochondria-rich cells increase in height, and protein synthesis (as suggested by rough endoplasmic reticulum proliferation) increases dramatically. No new cell types or alterations in morphology were noted among glands taken from either adult or juvenile snakes, even though the venom of each is quite distinct. In general, the glands of C. v. oreganus share structural similarities with those of crotalids and viperids previously described.     

Complex epidermal organs of Phascolion (Sipuncula): insights into the evolution of bimodal secretory cells in annelids

The epidermal organs of an undescribed Phascolion species from the Balearic Islands were investigated using SEM, TEM, LM, CLSM and μCT methods. We found axial receptor cells confirming the previously assumed sensory function of epidermal organs. Our analyses also revealed six types of secretory cells. Some secretory cells types are capable of secreting filamentous and amorphous secretion in two different ways simultaneously (bimodal secretion). The high diversity of cell types, the complex pattern of acinar units, and the absence of a common gland pore make epidermal organs of Phascolion unique amongst sipunculans (Phascolion type). Our reconstruction of the evolution of the epidermal organs of Sipuncula revealed that Phascolion‐type epidermal organs may have derived from either Golfingia‐, Sipunculus‐ or Phascolosoma‐type epidermal organs. The oldest known sipunculans were Golfingia‐like and had epidermal organs, which might resemble the architecture of the Golfingia‐type epidermal organs in extant taxa. Thus, it can be hypothesized that bimodal secretory cells (e.g. basophilic secretory cells) were part of the sipunculan ground pattern. Moreover, bimodal secretory cells of Phascolion look strikingly similar to those found in various annelid glands and thus might even be part of the ground pattern of stem species of Sipuncula + Pleistoannelida.     

The Secretory Structure of Hypericum perforatum and Its Relation to Hypericin Accumulation

Three different kinds of internal secretory structures, secretory cell globules (black dots), secretory cavities (translucent dots) and secretory ducts (translucent streaks) were observed in Hypericum perforatum L. The secretory cell globule, which occurred in flower, leaf and stem, consisted of a core of large secretory cells surrounded by two layers of flattened sheath cells. The secretory cavities were present throughout the lamina and the secretory ducts throughout the flower, both consisted of one or two layers of flattened cells walling an oil chamber or duct. Histochemical and fluorescence microscopy revealed that hypericin was accumulated in the secretory cell globules. Microscopic and ultrastmctural observation further demonstrated that hypericin was accumulated in the large central vacuole of the mature secretory cells. Numerous dictyosome, endoplasmic reticulum and small vacuoles were observed in the dense cytoplasm surrounded the large central vacuole. The process of hypericin accumulation in the secretory cells was preliminarily discussed.     

Survey of leaf anatomy,especially secretory structures,of tribeCaesalpinieae (Leguminosae,Caesalpinioideae)

We studied leaflet anatomy, emphasizing secretory structures, from herbarium specimens of 128 species of 44 genera of tribeCaesalpinieae, using clearings, resin sections, and scanning electron microscopy. These observations, combined with those from our three earlier papers, provide a survey of 210 species representing all genera. Seventy-three species had secretory structures: 21 had glands or gland-like trichomes, 40 had living mesophyll idioblasts, and nine had cavities (three species each had two different types). Five additional species, all inCercidium (Caesalpinia group), had paired or clustered large spheroidal, thick-walled, empty cells (veinlet idioblasts) interconnected by perforation plate-like gaps. Secretory structures have systematic significance at various taxonomic levels.     

Secretory Cavity Development and Its Relationship with the Accumulation of Essential Oil in Fruits of Citrus medica L. var. sarcodactylis （Noot.） Swingle

The developmental types of secretory cavities in Citrus remain controversial. The relationship between secretory cavity development and the accumulation of essential oil in fruits of Citrus species is also unknown. In order to develop better insights into these problems, histological, histochemical, and cytochemical methods were used to investigate secretory cavity development and the accumulation of essential oll at different developmental stages of fruits of Citrus medica L. var. sarcodactylis （Noot.） Swingle. The results indicate that the secretory cavity of the variety seemed to originate from an epidermal cell and a subepldermal cell. These two cells underwent successive divisions, resulting In the formation of two parts： （Ⅰ） a conical cap; and （Ⅱ） a globular gland. The formation of the lumen was schlzolysigenous. Regular changes in the size of vacuoles and the accumulation of essential oil were revealed during the process of secretory cavity development. In addition, when fruits were a light yellow or golden color, the structure of secretory cavities was well developed and the content of essential oil in a single fruit reached a maximum. It would be most appropriate to collect the fruit as a medicinal material at this time.     

Immunolocalization and histochemical evidence for the association of two different<Emphasis Type="Italic"> Arabidopsis</Emphasis> annexins with secretion during early seedling growth and development

Annexins are a multigene, multifunctional family of calcium-dependent, membrane-binding proteins found in animal and plant cells. In plants, annexins have been localized in the cytoplasm and at the cell periphery of highly secretory cell types, and in the tip region of polarly growing cells. Consequently, one proposed function for annexins in plant cells is participation in the Golgi-mediated secretion of new wall materials. In Arabidopsis, there are eight different annexin cDNAs, which share between 30% and 81% deduced amino acid sequence identity. We have used two monospecific Arabidopsis anti-annexin antibodies, raised against divergent 31-mer peptides from AnnAt1 and AnnAt2 and a previously characterized pea anti-annexin p35 antibody, for Western blot and immunolocalization studies in Arabidopsis. Western blot analyses of various Arabidopsis protein fractions showed that the two Arabidopsis antibodies are able to specifically recognize annexins in both soluble and membrane fractions. Immunofluorescence results with the three annexin antibodies show staining of secretory cells, especially at the cell periphery in developing sieve tubes, outer root cap cells, and in root hairs, consistent with previous results. In developmentally different stages some staining was also seen near the apical meristem, in some leaf cells, and in phloem-associated cells. Autoradiography following ³H-galactose incorporation was used to more clearly correlate active secretion of wall materials with the localization patterns of a specific individual annexin protein in the same cells at the same developmental stage. The results obtained in this study provide further support for the hypothesis that these two Arabidopsis annexins function in Golgi-mediated secretion during early seedling growth and development.     

Autophagy delivers misfolded secretory proteins accumulated in endoplasmic reticulum to vacuoles in the filamentous fungus Aspergillus oryzae

Autophagy is a conserved intracellular degradation process of eukaryotic cells. In filamentous fungi, although autophagy has been reported to have multiple physiological roles, it is not clear whether autophagy is involved in the degradation of misfolded proteins. Here, we investigated the role of autophagy in the degradation of misfolded secretory proteins accumulated in endoplasmic reticulum (ER) in the filamentous fungus Aspergillus oryzae. In late-phase cultures, a disulfide bond-deleted mutant of the secretory protein α-amylase AmyB fused with mDsRed that had accumulated in the ER was subsequently delivered to vacuoles, whereas wild-type AmyB-mDsRed was predominantly located at cell walls and septa. To examine the involvement of autophagy in the delivery of mutant AmyB to vacuoles, mutant AmyB-EGFP was expressed in an A. oryzae autophagy-deficient strain (ΔAoatg8). Microscopic examination revealed that the protein delivery to vacuoles did not occur in the absence of autophagic activity, with mutant AmyB-mDsRed forming large spherical structures surrounded by ER membrane. Hence, we conclude that autophagy is responsible for the delivery of misfolded secretory proteins accumulated in the ER to vacuoles for degradation during late-growth phase in A. oryzae. This is the first study to provide evidence that autophagy plays a role in the degradation of misfolded secretory proteins in filamentous fungi.     

Ontogenetic plate and plate field development in two chitons,Middendorffia andIschnochiton

Summary The larva ofIschnochiton gradually develops 6 transversal ridges and later a seventh one on its back. These intersegmental ridges are composed of two types of cells, one of them a goblet cell, as they are found in the girdle. Both types of cell seem to be responsible for cuticle secretion. The depressions between the ridges, called plate fields, are responsible for plate secretion. There are two different cell types. The marginal ones, which remain slender and tall, develop an enormous flat villus (here called a stragulum). Each plate field is tightly sealed against sea water by the stragulum. In the crystallization chamber formed by the stragulum, the first plate anlage is formed. In the later stages, the stragulum protects the growing plate margin. No periostracum is found. Most probably the cells developing the stragulum are responsible for the secretion of the tegmentum and in early development are the only secretory cells. The central cells of the plate field, which very soon flatten, then produce the second plate layer, the hypostracum. There is no secretion product in common to plate fields and intersegmental ridges. During the developmental period studied inIschnochiton (from hatching to 94 h) the number of cells of the individual plate field is not increased.The earliest anlage of the plates inMiddendorffia andIschnochiton is rodlike with straight margins and is not composed of several granules. Growth takes place at least inMiddendorffia by apposition of lime to the ends, posterior and, preferably, anterior margins. All plates are posttrochal.     

Hemoglobin switching in the salamanderPleurodeles waltlii

Summary In order to study the cellular distribution of larval and adult hemoglobins during larval development ofPleurodeles waltlii a double specific immunofluorescent labelling technique was developed.Rabbit antibodies specific for larval and adult hemoglobin components were prepared and conjugated with tetramethyl-rhodamine isothiocyanate for the anti-larval antibodies and fluorescein isothiocyanate for the anti-adult hemoglobin antibodies.Both simultaneous and sequential staining with the two types of fluorescent antibodies indicated that larval and adult hemoglobins were never observed within the same erythrocyte during development. The results provide evidence that two distinct cell populations exist, one synthesizing exclusively larval hemoglobins which is progressively replaced by the other one synthesizing exclusively adult hemoglobins. It remains to be determined if these two populations arise from two distinct types of stem cells (adult and larval) or from the same stem cell type.     

Structure and histochemistry of the extrafloral nectary ofAcacia terminalis (Salisb.) MacBride (Leguminosae,Mimosoideae)

Summary The extrafloral nectary ofAcacia terminalis is of the flat type and is located on the adaxial surface of the petiole of the bipinnate leaf. The secretory area is restricted to the base of the trough and no gaps or pores were detected by staining with vital dyes. Between the vascular bundles beneath the nectary and the surface cuticle there were three cell types. The cells of the flanking zone adjacent to the vascular bundles did not appear to be producing secretion whereas the cells of the glandular and secretory zones were secreting. The cells of the glandular zone were elongated whereas those of the surface secretory zone were spherical. Both had endoplasmic reticulum and Golgi bodies with secretory vesicles which were observed in close association with the plasmalemma. Secretion accumulated in the intercellular spaces of the glandular zone cells and forced the cells of the secretory zone apart. Symplastic contact was maintained in all cell types by plasmodesmata which were often associated with endoplasmic reticulum. Secretion accumulated beneath the cuticle which was distended but remained intact on the surface of the secretion.     

The Golgi apparatus in epidermal mucoid and ellipsoid-vacuolate cells ofAeolidia papillosa andCoryphella rufibranchialis (Nudibranchia)

Summary The epidermal cell layer of the apical end of the ceras was investigated in two species of aeolid nudibranchs. Based on cellular inclusions, mostly two cell types were found: mucoid and ellipsoid-vacuolate cells. Mucoid cells ofCoryphella rufibranchialis have large heterogeneous and fibrillar secretory granules whereas inAeolidia papillosa, the granules are homogeneous, but vary in electron density from one cell to another. Ellipsoid-vacuolate cells contained large quantities of small vacuoles with an included ellipsoidal structure. Both species contained very numerous ellipsoid-vacuolate cells. Secretory granules and ellipsoid-vacuoles appear to arise from the Golgi apparatus and these contents stain with PAS, suggesting a polysaccharide composition. Mucoid cells contained both secretory granules and ellipsoid-vacuoles which may arise from the same Golgi apparatus.     

Ultrastructural localization of egg-laying prohormone-related peptides in the atrial gland of Aplysia californica

The atrial gland is an exocrine organ that secretes into the oviduct of Aplysia californica and expresses three homologous genes belonging to the egglaying hormone gene family. Although post-translational processing of the egg-laying hormone precursor in the neuroendocrine bag cells has been examined in detail, relatively little is known about the post-translational processing of egg-laying hormone-related gene products in the atrial gland. A combination of morphologic techniques that included light-microscopic histology and immunocytochemistry, transmission electron microscopy, and immuno-electron microscopy were used to localize egg-laying hormone-related peptides in the atrial gland and to evaluate the characteristic morphology of their secretory cells. Results of these studies showed that there were at least three major types of secretory cells in the atrial gland (types 1–3). Significantly, of these three cell types, only type 1 was immunoreactive to antisera against egg-laying hormone-related precursor peptides. The immunoreactivity studies established that all three egg-laying hormone-related precursor genes are expressed in type-1 cells and indicated that the processing of these precursors also occurs within the secretory granules of this cell type. Evidence was also obtained that proteolytic processing of the egg-laying hormone-related precursors differed significantly from that observed in the bag cells. In contrast to the bag cells, the NH₂-terminal and COOH-terminal products of the egg-laying hormone-related precursors of the atrial gland were not sorted into different types of vesicles.     

Correlations between Accumulation of Dianthramides, Dianthalexin and Unknown Compounds, and Partial Resistance to Fusarium oxysporum f. sp. dianthi in Eleven Carnation Cultivars

The accumulation of dianthramides, dianthalexin and other phenolic compounds in carnation cells at different times before and after inoculation of plants with Fusarium oxysporum f. sp. dianthi, and the development of wilt symptoms were compared for eleven carnation cultivars. Untreated and water-treated controls contained only a little acetone-extractable phenolics. Upon inoculation with F. oxysporum f. sp dianthi, all cultivars accumulated the same range of phenolic compounds, including several dianthramides and dianthalexin, but in different proportions. The total amount of accumulated phenolic compounds per fresh material weight of the extracted stem segments could not be related to the disease resistance level of the different cultivars. However, the accumulation of dianthramides and dianthalexin in the cultivars was positively correlated to resistance, while the accumulation of two other, as yet unidentified, compounds was inversely related to resistance.