Changes of cell morphology and prolactin secretion induced by 2-Br- alpha-ergocryptine, estradiol, and thyrotropin-releasing hormone in rat anterior pituitary cells in culture

The secretion of prolactin in cultured pituitary cells was studied in correlation with the cellular changes induced by stimulatory or inhibitory agents. The techniques used in this study were: radioimmunoassay, immunocytochemistry, scanning (SEM) as well as transmission (TEM) electron microscopy. Prolactin secretion was stimulated by 17 beta-estradiol (10 nM) as well as thyrotropin- releasing hormone (TRH) (3 nM) and inhibited by 2-Br-alpha-ergocryptine (CB-154) (1 muM). The total prolactin (release and cell content) increased between 2 and 8 d of estradiol treatment, indicating an increase of both synthesis and release of prolactin. This finding was in agreement with TEM observations because, in estradiol-treated prolactin cells, the Golgi saccules were distended and Golgi elements were increased, thus indicating increased synthetic activity of these cells. The addition of TRH over a 4-h period resulted in a significant degranulation of prolactin cells. In contrast, prolactin secretory granules became accumulated in the cells after CB-154 treatment for a period ranging from 4 to 24 h. In agreement, light microscope immunocytochemistry showed an increased reaction for prolactin after short-term (< 24 h) incubation with CB-154. Because prolactin cells represent approximately 70% of the glandular cell population as revealed by immunocytochemistry, it was then possible to observe the changes of cell surface by SEM. In most cells, estradiol and TRH led to an increase in the number and prominence of microvilli and blebs, whereas CB-154 treatment resulted in a slightly decreased number of microvilli and an increased occurrence of membrane foldings. This report thus provides morphological evidence for the stimulatory effects of estradiol and TRH, and the inhibitory effects of CB-154 on prolactin secretion in pituitary cells in primary culture. These data, moreover, show that acute changes in secretory activity of prolactin-secreting cells are accompanied by marked changes of their morphological characteristics.     

Effects of colchicine and 2-Br-alpha-ergocryptine-methane-sulfonate (CB 154) on the release of prolactin and growth hormone by functional pituitary tumor cells in culture

The effects of colchicine and 2-Br-α-ergocryptine-methane-sulfonate (CB 154) on the release of prolactin and growth hormone have been studied in a clonal strain of rat pituitary tumor cells (GH₃) in monolayer culture. These cultures produce both prolactin and growth hormone and release both proteins spontaneously into the medium without storing them in large amounts. Immunological methods were used to measure both intracellular and extracellular concentrations of the hormones. Colchicine (5 × 10^?6 M for 3 hours) caused a 2- to 3-fold increase in intracellular concentrations of prolactin and growth hormone but, under basal conditions, had little or no measurable effect on the amounts of hormone accumulated in the medium during the course of the standard three hour treatment period. This latter finding evidently is due to a lag in the onset of drug action. Colchicine had little or no effect on accumulation of extracellular prolactin during the first two hours of treatment whereas such accumulation was depressed by over 60% during the third hour of treatment. Previous studies have shown that treatment of GH₃ cells with thyrotropin releasing hormone (TRH) and hydrocortisone (HC) increases both intra and extracellular levels of prolactin and growth hormone, respectively. In cultures treated with TRH (5 × 10^?8 M), colchicine (5 × 10^?6 M for 3 hours) increased intracellular prolactin by about 70% and decreased extracellular hormone by 10%. In cultures treated with HC (3 × 1O^?6 M), colchicine increased intracellular growth hormone by more than 100% and decreased medium concentrations of the hormone by 15%. Colchicine did not significantly alter total hormone (intracellular + extracellular) accumulation, cellular uptake of ³H-amino acids, or total cell protein synthesis. The synthetic ergot alkaloid, CB 154, (3.3 × 10^?6 M for 3 hours) caused an 80% increase in intracellular, and a nearly 50% decrease in extracellular, prolactin without affecting the accumulation of growth hormone, the uptake of ³H-labeled amino acids, or overall protein synthesis in the cultures. Elevation of medium potassium concentration from a basal value of 5.3 mM to 3–5 × 10^?2 M (by addition of KCl) decreased intracellular levels of prolactin by 85% and growth hormone by 55%. These effects of high potassium were blocked by colchicine and by CB 154. We conclude that colchicine, after a lag period of two hours, acts to inhibit the release of prolactin and growth hormone from GH₃ cells. By the end of three hours of treatment, this inhibition is over 60% complete in the case of prolactin. The qualitatively different effects of colchicine and CB 154 on prolactin and growth hormone release suggest that these two secretory blocking agents probably act on GH₃ cells by different mechanisms.     

Microtubule regulation of cell surface receptor topography during granulosa cell differentiation

Abstract. A possible role for cytoplasmic microtubules in modulating lectin binding site topography has been examined during the hormone-directed differentiation of rat ovarian granulosa cells in vitro. Indirect immunofluorescence staining with anti-tubulin antibodies indicates that undifferentiated cultured granulosa cells contain a network of microtubules which radiate from the cell center to the cell periphery. Cultures induced to differentiate by a three day treatment with 1 μg/ml prolactin exhibit a marginal distribution of microtubules and a centrally-located primary cilium. Prolactin enhances the incidence of granulosa cells containing a primary colium from 9% in undifferentiated cultures to 53% in hormone-treated cultures. The pattern of lectin binding site redistribution induced by Concanavalin A (Con A) is also modified by prolactin treatment. In contrast to undifferentiated cells, which randomly endocytose fluorescein Con A, granulosa cells exposed to prolactin respond to fluorescein Con A by forming central surface caps to a greater extent (75%) than undifferentiated controls (25%). Double label fluorescence microscopy and transmission electron microscopy on Con A labeled cells show that caps form at central cell surface sites which contain the primary cilium. Disruption of cytoplasmic microtubules by colchicine, in undifferentiated granulosa cells, results in the formation of cell surface caps upon Con A addition. These data suggest that cytoplasmic microtubules modulate the topography of lectin bindings sites which is subject to hormonal control during the in vitro differentiation of ovarian granulosa cells.     

Microtubule regulation of cell surface receptor topography during granulosa cell differentiation

A possible role for cytoplasmic microtubules in modulating lectin binding site topography has been examined during the hormone-directed differentiation of rat ovarian granulosa cells in vitro. Indirect immunofluorescence staining with anti-tubulin antibodies indicates that undifferentiated cultured granulosa cells contain a network of microtubules which radiate from the cell center to the cell periphery. Cultures induced to differentiate by a three day treatment with 1 microgram/ml prolactin exhibit a marginal distribution of microtubules and a centrally-located primary cilium. Prolactin enhances the incidence of granulosa cells containing a primary cilium from 9% in undifferentiated cultures to 53% in hormone-treated cultures. The pattern of lectin binding site redistribution induced by Concanavalin A (Con A) is also modified by prolactin treatment. In contrast to undifferentiated cells, which randomly endocytose fluorescein Con A, granulosa cells exposed to prolactin respond to fluorescein Con A by forming central surface caps to a greater extent (75%) than undifferentiated controls (25%). Double label fluorescence microscopy and transmission electron microscopy on Con A labeled cells show that caps form at central cell surface sites which contain the primary cilium. Disruption of cytoplasmic microtubules by colchicine, in undifferentiated granulosa cells, results in the formation of cell surface caps upon Con A addition. These data suggest that cytoplasmic microtubules modulate the topography of lectin bindings sites which is subject to hormonal control during the in vitro differentiation of ovarian granulosa cells.     

Changes in mitotic indices in roots of Vicia faba

Roots of Vicia faba were treated with solutions of colchicine or IAA or both. Mitotic indices and the frequencies of the different stages of mitosis were determined immediately after a three hour treatment or following a 24 hour period of recovery. Roots scored after treatment with colchicine for three hours showed several effects, none of which were reversed by simultaneous treatment with IAA. Treatment with IAA for three hours had little detectable effect on mitotic index (MI) on the frequencies of the various stages of mitosis. After a recovery period, following a three hour treatment, of 24 hours, colchicine treated roots showed a significant increase in their MI; this was due largely to an increase in the number of metaphases but it was also due in part to the presence of tetraploid cells in division. IAA treated roots revealed an inhibition of mitotic activity, which was most marked at 3.13–6.26×10^–4 M IAA. The results from roots treated with mixtures of colchicine and IAA for three hours and fixed 24 hours later showed: 1) the increase in MI induced by colchicine is reversed by IAA, the intensity of the reversal increasing with increasing concentrations of IAA; 2) reductions in the total numbers of cells in prophase or in metaphase occur after treatment with different concentrations of IAA; 3) IAA leads to a reduction in the number of tetraploid cells seen in division.It appears that colchicine induces a change in the pattern of mitotic activity 24 hours after the end of treatment and its effects are reversed by IAA. At 4.2×10^–4 M IAA a balance occurs between the opposing effects of colchicine and IAA and the MI is not significantly different from that of the controls. It is suggested that one result of a treatment with colchicine is a change in the level of growth factors in root meristems. This change, which appears to result in a temporary increase in MI is reversed by the addition of IAA. Thus one of the growth factors, the level of which has been affected, is replaceable by exogenous IAA.     

Morphological and cytochemical modifications induced by colchicine treatment in the duodenal epithelium of mouse

Summary S.W.R. adult male mice received one injection of colchicine and were killed at various time-intervals from 4 to 24 hours. Ultrastructural morphology and cytochemical activity of acid phosphatase, arylsulfatase and thiamine pyrophosphatase were recorded in the duodenal epithelium. Extensive damage was observed in the undifferentiated cells with formation of large heterophagosomes and in the absorptive cells with apparition of cytolysomes. The activity of the acid phosphatase and thiamine pyrophosphatase decreased after 6 hours of treatment. Moreover, the migration of the acid phosphatase content of the Golgi vesicles was restraint. These observations were related to the disruptive action of the alcaloid towards the microtubules.This work was supported by a grant from the Medical Research Council of Canada.Mr. Raymackers was a recipient of a studentship of the Medical Research Council of Canada. The authors are greatly indebted to Mr. Michel Couture and Pierre Magny for their skilful technical aid.     

Subcellular distribution of secretogranins I and II in GH3 rat tumoral prolactin (PRL) cells as revealed by electron microscopic immunocytochemistry

The GH3 rat pituitary cell line which secretes prolactin (PRL) is characterized by the paucity and small size of secretory granules. We looked for the presence, in these cells and in normal PRL cells, of two acidic tyrosine-sulfated proteins which are widely distributed in dense-core secretory granules of endocrine and neuronal cells, secretogranins I and II, using immunofluorescence and electron microscope immunoperoxidase techniques. Both secretogranins were detected in secretory granules of GH3 cells and of normal cells. Moreover, with our pre-embedding approach, secretogranins were localized within some RER cisternae and within all sacules of the Golgi stacks in both PRL cell models. A few small vesicles, large dilated vacuolar or multivesicular structures, and some lysosome-like structures were also immunoreactive. Double localization of secretogranins and PRL performed on GH3 cells by immunofluorescence indicated that all cells contained secretogranins I and II, whereas only 50-70% of the cells contained PRL. Moreover, in the case of hormone treatment known to increase the number of secretory granules, most if not all mature secretory granules were immunoreactive for secretogranins, whereas in certain cells some of the granules were apparently not immunoreactive for PRL. These immunocytochemical observations show that GH3 cells, which under normal conditions form only a small number of secretory granules, produce secretogranins and package them into these granules.     

Effects of colchicine on the ultrastructure of mouse taste buds

Summary Effect of colchicine on the ultrastructure of taste bud cells was studied in the mouse. In untreated mice microtubules were abundant throughout the entire cytoplasm of type-III cells, but only in the apical cytoplasm of type-I cells. After 2 h of colchicine treatment, no microtubules were observed in any taste bud cells; dense secretory granules in the apical cytoplasm of type-I cells mostly disappeared, and instead, numerous phagosomes appeared. It is suggested that colchicine causes an interruption of the transport of the secretory granules in type-I cells from the Golgi apparatus to the membrane of the apical surface, from which release occurs. In type-III cells, after 4 or 5 h of treatment, dense-cored vesicles scattered throughout the cytoplasm tended to increase in number; they were often observed to accumulate in the vicinity of the Golgi apparatus. Five hours after treatment with 5-hydroxy-l-tryptophan (5-HTP) following colchicine pretreatment, monoamine specific fluorescent cells and vesicles with highly electron-dense cores of type-III cells were still present. On the other hand, 5 h after 5-HTP treatment alone both fluorescent cells and vesicles with highly electron-dense cores had already disappeared. These observations suggest that the treatment with colchicine interrupts the transport of densecored vesicles of type-III cells to synaptic areas, in which those vesicles are presumed to discharge the neurotransmitter substance.     

Effect of colchicine on the intracellular transport of secretory proteins in rat liver parenchymal cells. Immunocytochemical observations

We studied the effects of colchicine on the intracellular transport of secretory proteins in rat liver parenchymal cells using the direct immunoenzyme technique. Livers were perfusion-fixed 0.5, 1, and 2 h after injection of colchicine. Vibratome sections of the fixed liver were stained using peroxidase-conjugated Fab' of anti-albumin or anti-fibrinogen. By light microscopy, reaction deposits showing albumin and fibrinogen were observed in the cytoplasmic granules of hepatocytes. Such stained granules decreased 30 min after injection, but later increased gradually and crowded in the cytoplasm. The Golgi complex stained for the proteins decreased after 30 min but increased in the juxtanuclear region after 60 min. The analysis of serial sections showed that colchicine severely disturbed the spatial relationship between the Golgi apparatus and the bile canaliculus. We obtained similar results by electron microscopy; a positive reaction for albumin and fibrinogen was observed in a small number of the cytoplasmic granules after 30 min. After 1 h of treatment, most of the Golgi complexes were fragmented and lost their stacked cisternae. However, they reappeared accompanied with vacuolated cisternae and secretory granules, which were partially stained for albumin and fibrinogen. After 2 h, the secretory granules positive for both proteins accumulated further. Some of them lined a long the plasma membrane, and others made a cluster in the cytoplasm. The profiles showing exocytosis were very rarely seen. These results showed that in the first 30 min, colchicine primarily disturbs partially the Golgi assembly but does not affect the post Golgi secretory pathway much. Later, the drug affects both the post Golgi pathway and the Golgi assembly, and it causes a marked accumulation of secretory granules.     

Fine morphological aspects of the secretory process in arterial smooth muscle cells. II. Role of microtubules

Preliminary results are presented regarding the role of the microtubular system in the secretory process in the rabbit aorta and pulmonary trunk smooth muscle cells. The results are based on electron microscopic findings of the effect of colchicine in vivo. On colchicine treatment an accumulation of secretory granules versus a vacuolar type of dilatation of the rough endoplasmic reticulum cisternae distinguishes two groups of smooth muscle cells. It is suggested that two secretory pathways operate in these cells, and that the cell microtubular status plays a key role in triggering one or the other pathway.     

Laminin induces formation of neurite-like processes and potentiates prolactin secretion by GH3 rat pituitary cells

Tumor-derived GH3 rat pituitary cell lines are widely utilized to study mechanisms of prolactin secretion and responsiveness to secretagogues. These cells served here as a model with which to study relationships between shape and function. When GH3 cells were routinely grown in serum-supplemented medium, they exhibited the polygonal phenotype of epithelial cells, with scarce secretory granules. In contrast, when seeded in a serum-free medium, they attached loosely and contained more secretory granules. In both cases, they released prolactin in a nonpolarized manner. We show in the present work that laminin extracted from Englebreth-Holm-Swarm (EHS) tumors was a potent attachment and spreading factor for GH3/B6 cells seeded in serum-free medium. Moreover, it induced the formation of neurite-like processes, which were increased in number and length by chronic treatment with a specific secretagogue, thyroliberin (TRH). These changes in cell shape were correlated with a potentiation of prolactin secretion, both basal and TRH-stimulated. Furthermore, using immunocytochemistry and electron microscopy, we revealed--at the dilated tip of processes--an accumulation not only of prolactin, but also of synaptophysin, a vesicle membrane marker, and of several organelles, such as secretory granules, smooth vesicles, dense bodies and mitochondria. The cytoplasmic processes contained long parallel bundles of microtubules and showed a strong immunoreactivity for beta 2-tubulin. In addition, we found immunocyto-chemical evidence for the presence of 200-k Da neurofilament protein in GH3/B6 cell processes as well as in neurites of cultured hypothalamic neurons. We conclude that, in GH3/B6 cells, laminin induced the differentiation of neurite-like processes, which were the site of polarized organelle transport and exhibited some neuronal markers.     

Ultrastructural and cytochemical studies of the effects of prolactin on the lateral prostate and the seminal vesicle of the castrated guinea pig

Summary Administration of ovine prolactin to castrated guinea pigs for 2 weeks induced hypertrophy of secretory cells in the lateral prostate when compared with the castrated controls. This was accompanied by an apparent increase in the number of profiles of granular endoplasmic reticulum and well developed Golgi complexes with dilated cisternae. An increase in the number of low-contrast electron-dense secretory granules was observed 4 weeks after prolactin treatment. In the seminal vesicle, dilatation and degranulation of granular endoplasmic reticulum and an apparent decrease in the number of secretory granules were observed 4 weeks after prolactin administration. Following castration and 2 weeks after prolactin treatment, thiamine pyrophosphatase (TPPase)-reaction product was mainly confined to 1–2 trans cisternae of the Golgi complexes in secretory cells of the lateral prostate and the seminal vesicle. In both glands, a reduction of TPPase activity was observed 2 weeks following prolactin administration, and the reaction product was totally absent after prolonged treatment for 4 weeks. The present study has provided morphological evidence that prolactin is capable of stimulating the secretory function of the lateral prostate while exerting some inhibitory effects on the seminal vesicle of the castrated guinea pig. In both glands, TPPase activity, and hence the process of glycosylation was inhibited after prolactin administration. The results from radioimmunoassay indicated that the action of prolactin on these glands could be a direct effect and not mediated through testosterone.     

Vinblastine-induced microtubular paracrystals in prolactin cells of anterior pituitary gland of lactating rats.

Vinblastine sulfate in physiological saline was injected directly into the pituitary glands of lactating rats. Injections were made through the ear canal using a syringe equipped with a 24-gauge needle. The animals were killed at 2, 4, or 6 hours after the injections. When the anterior pituitary glands were examined by electron microscopy, many microtubular paracrystalline deposits were seen in the prolactin and growth hormone cells. The usual cytoplasmic microtubules and microfilaments were not seen in the cytoplasm of these cells. Granular extrusion (exocytosis) was markedly depressed, and an accumulation of secretory granules was definitely observed in the prolactin cells after the administration of vinblastine.     

Membranes of mammary gland : XV. 5-Thio- -glucose decreases lactose content and inhibits secretory vesicle maturation in lactating rat mammary gland

Short-term administration of the glucose analog 5-thio- -glucose to primiparous lactating rats reduced mammary tissue lactose concentrations to half of control levels. Treatment with colchicine alone caused slight reductions in mammary tissue lactose content. These treatments did not alter the morphology or degree of development of rough endoplasmic reticulum or Golgi apparatus, but did cause alterations in secretory vesicles. In mammary tissue from untreated lactating animals, large, swollen secretory vesicles were abundant in apical regions of epithelial cells. After thioglucose administration secretory vesicles in the apical cytoplasm were smaller and were more densely packed with contents. While administration of colchicine alone caused accumulation of large numbers of nearly fully swollen vesicles, treatment with both colchicine and thioglucose induced accumulation of smaller, less fully developed secretory vesicles which contained morphologically recognizable casein micelles. Mammary tissue from late gestation rats was low in lactose; vesicles in this tissue resembled secretory vesicles in tissue from rats treated with thioglucose in that they were small and densely packed. These observations suggest that lactose, an osmoregulator in mammary gland, is transferred from Golgi apparatus to the apical cell surface within secretory vesicles. Lactose appears to be important for secretory vesicle maturation in mammary epithelial cells.     

Colchicine effects on meiosis in the male mouse

Antimitotic agents administered at the time of synapsis (leptotene/zygotene) have been shown to induce synaptic abnormalities visible during pachytene in the male mouse. The object of this study was to test the hypothesis that cells with relatively large amounts of colchicine-induced damage to the synaptonemal complex (SC) are eliminated from prophase whereas cells with relatively small amounts of SC damage proceed through to the end of prophase. Male mice were injected with tritiated thymidine to mark a cohort of spermatocytes at premeiotic S-phase for tracking through pachytene. Forty-eight hours later, when those cells were at leptotene/zygotene, colchicine was administered intratesticularly. Whole-mount SC spreads were made from animals sacrificed at various times following colchicine administration, and prepared for autoradiography. The marked cells were examined by light and electron microscopy and the kind and number of synaptic abnormalities were scored throughout pachytene. Colchicine-induced SC damage included single axial elements (univalents), together with partially synapsed and nonhomologously synapsed SCs. The amount of SC damage (amount and type per cell and frequency of cells with damage) scored at early pachytene exceeded by three- to fivefold the amount at late pachytene. This is consistent with spermatogenic cell loss from the seminiferous tubule via colchicine-induced destruction of Sertoli cell microtubules. The presence of spermatocytes with no more than four autosomal univalents at late pachytene indicates that some cells with low amounts of synaptic damage progress to the end of pachytene. The loss of the most severely damaged cells may represent a meiotic checkpoint at early pachytene in the male mouse. Received: 24 April 1996; in revised form: 29 August 1996 / Accepted: 11 March 1997     

The effects of tunicamycin on anterior pituitary hormone producing cells in the rat

An electron microscopic study was performed to clarify the effects of tunicamycin, a glycosylation inhibitor, on rat anterior pituitary cells. Tunicamycin (10, 50, and 100 micrograms/250 g B.W.) was intraperitoneally injected into rats, which were sacrificed 24 hrs later. Protein hormone producing GH and prolactin cells, and ACTH cells which are known to have a glycosylated precursor, showed no recognizable ultrastructural changes. TSH cells and gonadotrophs, both of which secrete glycoprotein hormones consisting of alpha and beta subunits, showed remarkable dilatation of the rough endoplasmic reticulum, and decreased numbers of secretory granules. These results suggest that the role of glycosylation in TSH cells and gonadotrophs may have a different biological significance from that in ACTH cells.     

Cytopathology of PC12 cells infected with Japanese encephalitis virus.

Infection of a clonal rat pheochromocytoma cell line, PC12, with Japanese encephalitis (JE) virus produced successively higher titers of virus in the culture fluid during the 72-h experimental period. In electron microscopical observation, JE virus entered PC12 cells by direct penetration through the plasma membrane at 2 min postinoculation (p.i.) and caused marked cellular hypertrophy and extensive proliferation of the cellular secretory system including rough endoplasmic reticulum (RER) and Golgi complexes starting 24 h p.i. The proliferating RER of the virally infected cells contained progeny virions and characteristic endoplasmic reticulum vesicles in its cisternae, and the proliferating Golgi complexes contained virions in their saccules. These findings indicated that the proliferation of the cellular secretory system occurred in association with viral replication and maturation in the system. Seventy-two hours p.i., the cellular secretory system of infected PC12 cells showed degenerative changes with vesiculation, disorganization, and dispersion of the Golgi complexes and fragmentation, focal cystic dilation, and dissolution of the RER in the same manner as those seen in the secretory system of JE-virus-infected neurons in the mouse brain. Thus, JE-virus-infected PC12 cells seem to be a suitable neurogenic cell line for the study of the pathogenic mechanism of JE virus. At the same time, the virally infected cells seem to offer an interesting cell model for the study of the morphogenesis of the cellular secretory system.     

Light and electron microscopic study of the degeneration and early regeneration of olfactory epithelium in the mouse.

Degeneration and early regeneration of olfactory epithelium from two strains of mice was studied at the light and electron microscopic levels from 12 hours to 3 days following nasal irrigation with 1% aqueous solution of zinc sulfate (ZnSO4) (a compound known to selectively damage olfactory epithelium). Distinct patterns of degeneration and stages of regeneration were evident following treatment. During the first 24 hours after treatment three progressive manifestations of the degenerative process were seen: (1) a relatively mild condition which was characterized by surface irregularities produced by cell protrusions, highly vacuolated cytoplasm, presence of large lysosome-like bodies and prominent intercellular spaces, (2) a more severe condition in which large areas of the epithelium were detached from the basement membrane cellular debris was present in the nasal chamber, and (3) a condition of total or near-total denudation of the epithelium of olfactory mucosa. The basal lamina was continuous and intact in most regions and the integrity of the subadjacent connective tissue was mostly well-preserved. Nerve bundles of the fila olfactoria were noted in varying degrees of degeneration during the course of the experiment. The most advanced neural degeneration was seen 24 to 72 hours after treatment. Onset of regeneration was suggested by the appearance of a simple squamous layer of cells above the basement membrane 48 to 72 hours after treatment. In addition to the simple epithelium a stratified epithelium consisting of two to four cell layers was also observed at this time. Glandular cells, containing secretory granules identical to those in Bowman's glandular cells, were noted in an apparent process of migration from the lamina propria into the the stratified epithelial layer. The last mentioned observation supports the proposition that new supportive epithelial cells originate from cells of Bowman's gland.     

鸭呼肠孤病毒强毒株致鸭胚原代成纤维细胞凋亡的研究

通过光镜、电镜、DNA Ladder法、流式细胞术、荧光染色对鸭呼肠孤病毒(DRV)诱导鸭胚原代成纤维细胞(DEF)凋亡情况进行检测.结果显示,光镜可见细胞形态学上出现细胞皱缩,染色质浓染边移;电镜观察到细胞胞浆浓缩,细胞核染色质凝聚、部分形成凋亡小体;荧光染色结果显示,在感染后24h有激发绿色荧光的凋亡细胞出现,随着时间的推移,激发红色荧光的死亡细胞数量增多;DNA Ladder检测到感染后24～144h的DNA样品呈梯形条带;流式细胞术于感染后24h检测到凋亡细胞,其数量在72～96h达到高峰,144h开始下降.研究结果表明,DRV在DEF增殖的过程中具有诱导宿主细胞凋亡的作用.     

Cytochemical localization of calcium and Ca2+,Mg2(+)-adenosine triphosphatase in colchicine-altered rat incisor ameloblasts

Colchicine is known to affect secretory, transport, and degradative functions of ameloblasts. The effects of colchicine on membrane-associated calcium and Ca2+,Mg2(+)-ATPase in secretory and maturation ameloblasts were investigated cytochemically. The pyroantimonate (PPA) method was used for localizing calcium and a modified Wachstein-Meisel medium was used to localize Ca2+,Mg2(+)-ATPase. Sections representing secretory and early maturation stages were examined by transmission electron microscopy. Morphological changes induced by colchicine included dislocated organelles and other well-established reactions to such anti-microtubule drugs. Calcium pyroantimonate (Ca-PA) deposits in most ameloblast types were markedly reduced, with the greater reduction occurring in those cells more severely altered morphologically. However, the cell membranes of both control and experimental smooth-ended maturation ameloblasts were essentially devoid of Ca-PA. The normal distribution and intensity of Ca2+,Mg2(+)-ATPase was not affected by colchicine. Because the observed reduction of membrane-associated calcium is apparently not mediated by Ca2+,Mg2(+)-ATPase in this case, other aspects of the calcium regulating system of ameloblasts are apparently targeted by colchicine.