Alteraciones del ciclo circadiano en las enfermedades psiquiátricas: papel sincronizador de la melatonina en el ciclo sueño-vigilia y la polaridad neuronal / Circadian cycle alterations in psychiatric diseases: melatonin role as a synchronizer of sleep-awake cycle and the neuronal polarity

Circadian rhythms are oscillations of physiological functions. The period of their oscillation is about 24 h, and can be synchronized by environmental periodic signals as night-day cycle. The endogenous periodical changes depend on various structural elements of the circadian system which consists of the effectors, the secondary oscillators, the synchronizers and the circadian pacemaker. In mammalian species, the physiological function better understood respect their oscillation pattern are the synthesis and release of several hormones (i.e. cortisol and melatonin), the body temperature, the sleep-awake cycle, the locomotive activity, cell proliferation, neuronal activity among other rhythms. The Suprachiasmatic nucleus is the main circadian pacemarker in mammals; its oscillation keeps the circadian system synchronized particularly with respect to the environment photo period. When light reaches the pigment melanopsin in ganglionar neurons in the retina, the photoperiod signal is sent to Suprachiasmatic nucleus, and its postsinaptic neurons distributes the temporal signal to pheripheral oscillators by nervous or humoral pathways. Among the oscillators, the pineal gland is a peripheral one modulated by Suprachiasmatic nucleus. At night, the indolamine melatonin is synthesized and released from pinealocytes, and reaches other peripheral oscillators. Melatonin interacts with membrane receptors on Suprachiasmatic nucleus pacemarker neurons, reinforcing the signal of the photoperiod. In mammals, exogenous melatonin synchronizes several circadian rhythms including locomotive activity and melatonin release. When this indolamine is applied directly into the Suprachiasmatic nucleus, it produces a phase advance of the endogenous melatonin peak and increases the amplitude of the oscillation. In humans, melatonin effect on the circadian system is evident because it changes the circadian rhythms phase in subjects with advanced sleep-phase syndrome, night workers or blind people. Also it reduces jet lag symptoms enhancing sleep quality and reseting the circadian system to local time. Melatonin effects on circadian rhythms indicate their role as a chronobiotic, since decreased daily melatonin levels that occur with age and in neuropsychiatric disorders are associated with disturbances in the sleep-awake cycle. In particular, it has been described that Alzheimer's disease patients have disturbed sleep-awake cycle and have decreased serum melatonin levels. Sleep disorders in Alzheimer's disease patients decrease when they are treated with melatonin. Moreover, sleep disturbances have been observed in bipolar disorder patients and often precede relapses of insomnia-associated mania and hypersomnia-associated depression. These disturbances are linked to delayed- and advanced- phases of circadian rhythms or arrhythmia; therefore, it has been suggested that bipolar disorder patients could be treated with light and dark therapy. In depressed patients, the levels of melatonin are low throughout the 24 hour period and have a delayed onset of the indolamine concentration and showed an advance of its peak. Schizophrenic patients have decreased levels in the plasmatic melatonin in both phases of the light-dark cycle. Melatonin administration to these patients increases their sleep efficiency. In addition, melatonin acts as a neuroprotector because of its potent antioxidant action and through its cytoskeletal modulation properties. In neurodegenerative animal models, its protector effect has been observed using okadaic acid. This neurotoxin is employed for reproducing cytoskeletal damage in neurons and increased oxidative stress levels, which are molecular events similar to those that occur in Alzheimer's disease. In N1E-115 cell cultures incubated with okadaic acid, the administration of melatonin diminishes hyperphosphorylated tau and oxidative stress levels, and prevents the neurocytoskeletal damage caused by the neurotoxin. Although it is known that melatonin plays a key role in the circadian rhythms entrainment, little is known about its synchronizing effects at molecular and structural level. In algae, it has been observed a link between morphological changes and the light-dark cycle and it is known that shape is determinated by the cytoskeletal structure. In particular, the alga Euglena gracilis changes its shape two times per day under the effect of a daily light-dark cycle. This alga has a long shape when there is a higher photosynthetic capacity at the half period of the day; on the contrary, it showed a rounded shape at the end of 24 h cycle. Also, the influence of the cell shape changes on the photosynthetic reactions was investigated by altering them with drugs that disrupt the cytoskeletal structure as cytochalasin B and colchicine. Both inhibitors blocked the rhythmic shape changes and the photo-synthetic rhythm. Moreover, there are some reports about cytoskeletal changes in plants targeted by circadian rhythms. Guarda cells of Vicia faba L. showed a diurnal cycle on the alpha and beta tubulin levels. In addition, it has been proposed that melatonin synchronizes different body rhythms through cytoskeletal rearrangements. In culture cells, nanomolar melatonin concentrations cause an increase in both the polimerization rate and microtubule formation through calmodulin antagonism. A cyclic pattern produced by melatonin in the actin microfilament organization has been demonstrated in canine kidney cells. Cyclic incubation of MDCK cells with nanomolar concentrations of melatonin, resembling the cyclic pattern of secretion and release to plasma produces a microfilament reorganization and the formation of domes. Studies in animals are controvertial regarding if the amount of microtubules in different tissues varies cyclically. In rats and baboons, melatonin administration or exposure of rats to darkness induced an increased number of microtubules in the pineal gland. However, in the hypothalamus, the exposure of rats to light resulted in an increase in the microtubular protein content. Similarly, (X-tubulin mRNA was augmented during the light phase in the hypothalamus, hippocampus and cortex. By contrast, in rats maintained in constant darkness, a decreased level in the tubulin content was observed in the visual cortex. Additional information on cycle variations observed in cytoskeletal molecules indicated that beta actin mRNA levels are lower during the day in the hippocampus and cortex. But no change was observed in actin protein levels in the cerebral cortex. However, increased levels of actin and its mRNA were observed in the hypothalamus. Exogenous melatonin administration at onset of night decreased the amount of actin in the hypothalamus, while the actin mRNA levels decreased when the administration was realized in the morning. In this review we will describe the synchronizer role of melatonin in the sleep-awake cycle and in the organization of cytoskeletal proteins and their mRNAs. Also, we will describe alterations in the melatonin secretion rhythm associated with a neuronal cytoskeleton disorganization in the neuropsychiatric diseases such as Alzheimer, depression, bipolar disorder and schizophrenia.

Los ritmos circadianos son patrones de oscilación con un periodo cercano a 24h que se observan en los procesos fisiológicos. En los mamíferos se han descrito funciones biológicas con regulación circádica tal como el ciclo sueño-vigilia. La administración de la melatonina, una indolamina secretada por la glándula pineal, sincroniza los ritmos circadianos. En los humanos, este efecto se ha estudiado en sujetos con síndrome de <

TiO2 nanotube stimulate chondrogenic differentiation of limb mesenchymal cells by modulating focal activity

Vertically aligned, laterally spaced nanoscale titanium nanotubes were grown on a titanium surface by anodization, and the growth of chondroprogenitors on the resulting surfaces was investigated. Surfaces bearing nanotubes of 70 to 100 nm in diameter were found to trigger the morphological transition to a cortical actin pattern and rounded cell shape (both indicative of chondrocytic differentiation), as well as the up-regulation of type II collagen and integrin beta4 protein expression through the down-regulation of Erk activity. Inhibition of Erk signaling reduced stress fiber formation and induced the transition to the cortical actin pattern in cells cultured on 30-nm-diameter nanotubes, which maintained their fibroblastoid morphologies in the absence of Erk inhibition. Collectively, these results indicate that a titanium-based nanotube surface can support chondrocytic functions among chondroprogenitors, and may therefore be useful for future cartilaginous applications.

Molecular imaging of membrane proteins and microfilaments using atomic force microscopy

Atomic force microscopy (AFM) is an emerging technique for a variety of uses involving the analysis of cells. AFM is widely applied to obtain information about both cellular structural and subcellular events. In particular, a variety of investigations into membrane proteins and microfilaments were performed with AFM. Here, we introduce applications of AFM to molecular imaging of membrane proteins, and various approaches for observation and identification of intracellular microfilaments at the molecular level. These approaches can contribute to many applications of AFM in cell imaging.

Avaliação de parametros funcionais e morfologicos da ação da angiotensina II sobre segmentos de tubulos proximais isolados e conservados em soluções de euro-collins e belzer / Funcional and morphological behavior of preserved PCT during treatment using angiotensin II and Losartan

Introdução: A qualidade da conservação obtida em um órgão e a duração do armazenamento seguro são dependentes do tipo de solução e tempo de conservação, estando sujeitas a intercorrências imprevisíveis. Em virtude disso, o parênquima renal e, particularmente, o túbulo contornado proximal (TCP), local onde ocorre necrose tubular aguda (NTA), são susceptíveis às mudanças na homeostase funcional do órgão, durante a conservação e durante a reperfusão pós-implante. O TCP reabsorve cerca de 70% de Na+ e H2O filtrados; sua membrana apical tem células especializadas com microvilosidades para aumentar a área de transporte unidirecional de solutos do lúmen para o sangue. O principal componente estrutural da bordadura em escova, filamentos de F-actina, interage com uma variedade de proteínas transmembranas, inclusive moléculas transportadoras de íons. Esta função é regulada por parâmetros fisiológicos e hormonais, sendo a Angiotensina II (AII) uma das principais envolvidas com o citoesqueleto de actina e proteínas associadas, mediada principalmente por seus receptores AT1. Métodos: Neste trabalho, túbulos isolados de fragmentos frescos e de fragmentos conservados por períodos de 1 e 24h nas soluções de Euro - Collins (ECO) e de Belzer (UW) foram tratados com AII, losartan e AII+losartan, para medida do volume de absorção de fluido (Jv=nl. min_1. mm-1 (microperfusão in vitro), e medida da intensidade (média) de pixel da fluorescência dos receptores AT1 e do citoesqueleto de actina (imunoistoquímica). Resultados: a) Demonstrou-se que AII (10-12M) estimula absorção fluida (Jv), que é diminuída com losartan (10-6M) em TCP frescos e também em TCP conservados em soluções...

Introduction: Depending on the preservation quality and duration, considering the solution composition and the preservation method, unexpected intercurrences may occur. In consequence, the renal parenchyma and particularly the proximal convoluted tubule (PCT), site where the acute tubular necrosis (NTA) occurs, are susceptible to changes in the organ functional homeostasis during preservation in the post implantation reperfusion period. The PCT reabsorbs approximately 70% of filtered Na+ and H2O, and its apical membrane has specialized cells with microvilli to increase the area of unidirectional solute transport from the lumen to the blood. The major structural component of brush border microvilli is the filamentous F-actin, which has been shown to interact with a variety of transmembrane proteins, including ion transport molecules. This function is regulated by physiological and hormonal parameters, such as angiotensin II (AII) interacting with actin cytoskeleton and associated proteins, mediated mainly through type I receptors (AT1R). After via different signaling pathways having been triggered, the varied functions are started, including fluid absorption (Jv), which is directly related to the actin cytoskeleton. Methods: Fresh (no preserved) and tubules preserved for 1h and 24hrs in Euro-Collins (EC) and Belzer (UW) solutions, treated with AII, losartan and AII+losartan, evaluated by "in vitro" microperfusion technique (fluid absorption (Jv=nl. min-1. mm-1), and by immunohistochemical technique (AT1 receptor measurement and actin cytoskeleton). Results: a) Our results showed that AII (10-12M) (physiological concentration) stimulates...

Organized Filaments in the adhesive system of Macrostomum tuba Graff, 1882 ( Platyhelminthes, Macrostomida)

The adhesive organs or “duo-gland adhesive organs” of platyhelminths are formed by a specialized epithelialcell and extensions of two gland cells. These organs are used for temporary fixation of the organism tosurfaces in aquatic habitats. The mechanisms involved in adhesion to and release from a given surfacedepend on secretions produced by the glands; less is known about the involvement of cytoplasmic filamentsin the anchoring cell itself. In this study, we examined the structure of the adhesive organs present in thetail plate of Macrostomum tuba Graff, 1882 (Platyhelminthes, Macrostomida), a freshwater, free-livingflatworm. Scanning and transmission electron microscopy allowed elucidation of the three-dimensionalorganization of the adhesive system, especially of the microvilli that formed the outer collar (or papilla),which was endowed with a fibrous core. Electrical stimulation caused the flatworms to extend their papillaeabove the ciliated surface. The use of tannin- and diamine-containing fixatives showed that the filamentousarray contained tonofilaments and actin filaments. Tonofilaments concentrate in the axis of each microvillus;actin filaments, about 7-8 nm thick, spread out towards the periphery. Scanning images demonstrated thefinger-like shape of the papillae, about 7-8 ìm high, with a terminal opening. Microvilli followed a straightcourse along the surface.

The effects of arsenic trioxide on the cell cycle and microfilament cytoskeleton in human nasopharyngeal carcinoma cell line / 中国耳鼻咽喉头颈外科

OBJECTIVE To evaluate the effects of arsenic trioxide (As2O3) on the cell cycle and microfilament cytoskeleton in human nasopharyngeal carcinoma cell line(CNE1),as well as possible mechanisms. METHODS The variation of cell cycle and microfilament cytoskeleton in CNE1 were observed using the flow cytometry (FCM),the laser scanning confocal microscopy(LSCM)and technology of fluorescence.RESULTS FCM showed that the proportion of G1 phase cells significantly increased in cells exposed to 2 and 4?mol/L As2O3(P

Increased Microfilaments in Hepatocytes and Biliary Ductular Cells in Cholestatic Liver Diseases

To assess the extent of microfilaments in cholestatic liver diseases we examined the cytoplasmic microfilaments in intrahepatic and extrahepatic cholestasis in man by electron microscopy. Study subjects were two patients with drug-induced intrahepatic cholestasis, three patients with intrahepatic cholestasis due to viral hepatitis, four patients with extrahepatic cholestasis due to stones of the common bile duct and two patients with primary biliary cirrhosis. Two biopsied specimens from patients without clinical or histological evidence of liver disease served as noncholestatic controls. The microfilaments in hepatocytes and biliary ductular cells were significantly increased in cholestasis compared with those in non-cholestatic controls. Well developed bundles of microfilaments were noted around the pericanalicular ectoplasm and seemed to be parallel to plasma membrane of the hepatocytes in cholestasis. In cholestasis, there were increased bundles of microfilaments around the periluminal region, lateral cell wall, and nucleus of biliary ductular cells. Two patterns of microfilaments bundles (fine microfilamentous network and spindle-shaped dense or clusters of microfilaments) were associated with cholestasis. The clustered form of microfilaments also seemed to be clearly associated with intracytoplasmic vacuoles containing bile salts. In conclusion, the increase of microfilaments in hepatocytes and biliary ductular cells may be the consequence of various forms of cholestasis. Further studies are needed to clarify the functional significance of increased microfilaments in cholestasis.

Vesicular transport as a new paradigm in short-term regulation of transepithelial transport

The vectorial transepithelial transport of water and electrolytes in the renal epithelium is achieved by the polarized distribution of various transport proteins in the apical and basolateral membrane. The short-term regulation of transepithelial transport has been traditionally thought to be mediated by kinetic alterations of transporter without changing the number of transporters. However, a growing body of recent evidence supports the possibility that the stimulus-dependent recycling of transporter-carrying vesicles can alter the abundance of transporters in the plasma membrane in parallel changes in transepithelial transport functions. The abundance of transporters in the plasma membrane is determined by net balance between stimulus-dependent exocytic insertion of transporters into and endocytic retrieval of them from the plasma membrane. The vesicular recycling occurs along the tracts of the actin microfilaments and microtubules with associated motors. This review is to highlight the importance of vesicular transport in the short-term regulatory process of transepithelial transport in the renal epithelium. In the short-term regulation of many other renal transporters, vesicular transport is likely to be also involved. Thus, vesicular transport is now emerged as a wide-spread general regulatory mechanism involved in short-term regulation of renal functions.

The Effect of Local Anesthetics on Neurulation of Early Chick Embryos

Chick embryos have been used widely as model systems for studies in experimental embryology and teratology. Especially early chick embryos are very useful for studies of neural tube defects. Local anesthetics such as lidocaine, tetracaine and ketamine are known to inhibit the formation of microfilaments and cause neural tube defects. We made a try to figure out the effect of local anesthetics on neurulation in early chick embryo, which is stage 8 by Hamburger & Hamiliton classification. After 6~9 hours incubation in media treated with above local anesthetics, we observed the effects of these agents by LM and EM. Stage 8 chick embryos responded to local anesthetics in a dose related manner. Tetracaine showed most potent effects. Neural folds failed to make a contact, so the neural tube was left open. In high dosage, the case of no growth or little growth of embryo was also observed. On scanning electron microscopic examination, the surface of neuroepithelium was flattened in embryos treated with ketamines. On transmission electron microscopic examination. neuroepithelium of embryos cultured in ketamine showed the decrease of microfilaments and less conspicuous feature of mitochondria and rough ER(endoplasmic reticulum)'s.

Effect of LPS on phagocytosis of rat Kupffer cells in vitro / 中国病理生理杂志

AIM: To investigate the effect of LPS on phagocytosis of Kupffer cells in vitro. METHODS: Isolated Kupffer cells were treated with LPS in vitro . The phagocytosis, microfilament, microtubules and apoptosis of Kupffer cells were determined by the beads phagocytosis test, fluorescence staining, fluorometry and flow cytometric analysis. RESULTS: LPS enhances the phagocytosis, actin content, microtubules fluorescence density of Kupffer cells in vitro , while at a large dose or for a long time, it lessened the phagocytosis increasing or phagocytosis, inhibites the microfilament and microtubules expression, and induced apoptosis. CONCLUSION: LPS enhances the phagocytosis of Kupffer cells in vitro , but in large amount, it inhibites the phagocytosis of Kupffer cells, which is probably related to LPS -induced microfilament, microtubules expression changes and apoptosis in Kupffer cells.

Expression of CD2AP and F-actin in rats with puromycin aminonucleoside nephrosis / 中国病理生理杂志

AIM: To investigate the role of CD2AP and F-actin in the pathogenesis of puromycin aminonucleoside nephrosis in rats. METHODS: Puromycin aminonucleoside nephrosis was induced by single intraperitoneal injection of puromycin aminonucleoside (PAN). Renal tissues were studied at 3, 7, 10 and 20 days after PAN injection by means of immunohistochemistry, RT-PCR, Western blotting and fluorescence. RESULTS: At day 3, CD2AP expression in podocytes began to decrease, and significantly decreased at day 7 and 10 (P

Effects of cytomegalovirus on expression of ?-actin mRNA and microfilaments in human embryo fibroblasts / 中国病理生理杂志

AIM: To investigate the infection of huma n embryo fibroblasts (HF) with CMV as well as the effect of CMV on ?-actin and mi crofilaments. METHODS: RT-PCR assay was used to detect the mRNA expression of CMV immediate early (IE) gene, ?-actin and GAPDH genes in HF cells infected wit h CMV. The morphological changes and microfilaments in infected cells were obser ved with transmission electron microscopy. RESULTS: The morphology of HF cells infected with CMV changed si gnificantly from fusiform shape to round shape. The mRNA expression of CMV immed iate early gene was detected. The increase in mRNA level of IE gene was parallel with the infected titer of CMV. However, t he expression level of ?-actin mRNA in HF cells infected with CMV was decreased compared with the uninfected cells, while the expression of GAPDH mRNA did not change. CMV particles were observed in the cells by electron microscope. Microfi laments were found ruptured and shortened after the infection of CMV. CONCLUSIONS: CMV was able to infect human embryo fibroblasts and replicated in the cells. Also the CMV infection affected the expressi on of ?-actin mRNA and the microfilaments.

Effects of colon carcinoma cells on stimulating canalization of human lymph endothelial cells in vitro / 解放军医学杂志

Objective To explore the effects of colon carcinoma cells on stimulating canalization of human lymphatic endothelial cells(hLECs)in vitro.Methods hLECs in experiment group were cultured with the supernatant of colon carcinoma cell SW480,and they were cultured in endothelial culture medium in control group.The difference of the 2 groups in the ability of canalization was observed,the changes in cytoskeleton and the expression of Prox1 were detected by immunofluorescence assay,and the expression of integrin ?9 was determined by Western blotting.Results In comparison to the control group,hLECs in experiment group showed stronger ability of canalization,as a copious net-structure appeared on day 7 of cultivation,and the typical tube-structure formed finally on day 14.The number of tube-structure,including lymphatic branches,were greater in hLECs of experiment group(2.93?0.56)than control group(1.56?0.26)from day 6 on(P

STATE OF ACTIN POLYMERIZATION IN B LYMPHOCYTES FROM DIFFERENT DEVELOPMENT STAGES OF BURSA OF FABRICIUS OF PEKING DUCK / 解剖学报

In this study we have examined the state of actin polymerization in B cells from different development stages in bursa of Fabricius of Peking duck by indirect immunofluorescence technique using antibody against actin. In B cells from bursa of Fabricius of 26th day embryo and 3th week after hatching, the polymeric actin was the main form in the actin pool. In B cells from bursa of Fabricius of 12th week after hatching, however, the monomeric actin was the main form, which might be a result of the shift from the polymeric actin pool to the monomeric pool. We concluded that the state of actin polymerization might be an important factor in the cellular functions of B cells of bursa of Fabricius.

ULTRASTRUCTURAL FEATURES OF THE POSTNATAL DEVELOPMENT OF THE ECTOPLASMIC SPECIALIZATION IN RAT SERTOLI CELLS / 解剖学报

The postnatal development of the ectoplasmic specialization (ES) of Sertoli cells was investigated in the rats from 1st to 8th week after birth and the adult by electron microscopy. At the end of the 1st week. ES is in the beginning of its formation. Some high electron dense materials accumulate in the submembranous regions and short cisternae of endoplasmic reticulum align in these zones. At the end of the 2nd week the short cisternae are fused each other and straight microfilaments bundles appear and are sandwiched between the flattened cisternae and cell membrane. The density of microfilaments is increased in the 3th week and numerous tight junctions appear between adjacent Sertoli cells. The ES is gradually completed in its structure until the 5th week and fully extend and circumscribed around the base of Sertoli cells. Thereafter, there is no more morphological changes. We conclude that the first 5 weeks after birth is the important period for the development of ES of Sertoli cell as well as the blood-testis barrier in rats.