Mitofusins modulate the increase in mitochondrial length, bioenergetics and secretory phenotype in therapy-induced senescent melanoma cells.

Cellular senescence is an endpoint of chemotherapy, and targeted therapies in melanoma and the senescence-associated secretory phenotype (SASP) can affect tumor growth and microenvironment, influencing treatment outcomes. Metabolic interventions can modulate the SASP, and an enhanced mitochondrial energy metabolism supports resistance to therapy in melanoma cells. Herein, we assessed the mitochondrial function of therapy-induced senescent melanoma cells obtained after exposing the cells to temozolomide (TMZ), a methylating chemotherapeutic agent. Senescence induction in melanoma was accompanied by a substantial increase in mitochondrial basal, ATP-linked, and maximum respiration rates and in coupling efficiency, spare respiratory capacity, and respiratory control ratio. Further examinations revealed an increase in mitochondrial mass and length. Alterations in mitochondrial function and morphology were confirmed in isolated senescent cells, obtained by cell-size sorting. An increase in mitofusin 1 and 2 (MFN1 and 2) expression and levels was observed in senescent cells, pointing to alterations in mitochondrial fusion. Silencing mitofusin expression with short hairpin RNA (shRNA) prevented the increase in mitochondrial length, oxygen consumption rate and secretion of interleukin 6 (IL-6), a component of the SASP, in melanoma senescent cells. Our results represent the first in-depth study of mitochondrial function in therapy-induced senescence in melanoma. They indicate that senescence increases mitochondrial mass, length and energy metabolism; and highlight mitochondria as potential pharmacological targets to modulate senescence and the SASP.

Ultrastructural features of aberrant glial cells isolated from the spinal cord of paralytic rats expressing the amyotrophic lateral sclerosis-linked SOD1G93A mutation.

In the rat model of amyotrophic lateral sclerosis expressing the G93A superoxide dismutase-1 mutation, motor neuron death and rapid paralysis progression are associated with the emergence of a population of aberrant glial cells (AbAs) that proliferate in the degenerating spinal cord. Targeting of AbAs with anti-neoplasic drugs reduced paralysis progression, suggesting a pathogenic potential contribution of these cells accelerating paralysis progression. In the present study, analyze the cellular and ultrastructural features of AbAs following their isolation and establishment in culture during several passages. We found that AbAs exhibit permanent loss of contact inhibition, absence of intermediate filaments and abundance of microtubules, together with an important production of extracellular matrix components. Remarkably, AbAs also exhibited exacerbated ER stress together with a significant abundance of lipid droplets, as well as autophagic and secretory vesicles, all characteristic features of cellular stress and inflammatory activation. Taken together, the present data show AbA cells as a unique aberrant phenotype for a glial cell that might explain their pathogenic and neurotoxic effects.

Astrocyte Dysfunction in Developmental Neurometabolic Diseases.

Astrocytes play crucial roles in maintaining brain homeostasis and in orchestrating neural development, all through tightly coordinated steps that cooperate to maintain the balance needed for normal development. Here, we review the alterations in astrocyte functions that contribute to a variety of developmental neurometabolic disorders and provide additional data on the predominant role of astrocyte dysfunction in the neurometabolic neurodegenerative disease glutaric acidemia type I. Finally, we describe some of the therapeutical approaches directed to neurometabolic diseases and discuss if astrocytes can be possible therapeutic targets for treating these disorders.

Initial studies on mechanism of action and cell death of active N-oxide-containing heterocycles in Trypanosoma cruzi epimastigotes in vitro.

Chagas disease, endemic in 21 countries across Latin America, kills more people in the region each year than any other parasite-borne disease. Therapeutic options have problems ranging from toxicity, poor efficacy, drug resistance and high cost. Thus, cheaper and less toxic treatments are necessary. From our in-house chemical library of agents against Trypanosoma cruzi the most relevant N-oxide-containing heterocycles were selected for mode of action and type of death studies. Also included in these studies were two active nitrofuranes. Epimastigotes of T. cruzi were used as the biological model in this study. The metabolic profile was studied by 1H NMR in association with the MTT assay. Excreted catabolites data, using 1H NMR spectroscopy, showed that most of the studied N-oxides were capable of decreasing both the release of succinate and acetate shedding, the compounds therefore possibly acting on mitochondria. Only quinoxalines and the nitrofurane Nf1 showed significant mitochondrial dehydrogenase inhibitions, but with different dose-time profiles. In the particular case of quinoxaline Qx2 the glucose uptake study revealed that the integrity of some pathways into the glycosome could be affected. Optic, fluorescence (TUNEL and propidium iodide) and transmission electron microscopy (TEM) were employed for type of death studies. These studies were complemented with 1H NMR to visualize mobile lipids. At low concentrations none of the selected compounds showed a positive TUNEL assay. However, both quinoxalines, one furoxan and one benzofuroxan showed a necrotic effect at high concentrations. Curiously, one furoxan, Fx1, one benzofuroxan, Bfx1, and one nitrofurane, Nf1, caused a particular phenotype, with a big cytoplasmatic vacuole being observed while the parasite was still alive. Studies of TEM and employing a protease inhibitor (3-methyladenine) suggested an autophagic phenotype for Bfx1 and Nf1 and a 'BigEye' phenotype for Fx1.

Brain atlas of the annual Garcialebias charrua fish.

Annual fish have become attractive study models for a wide range of disciplines, including neurobiology. These fish have developed different survival strategies. As a result, their nervous system is under considerable selective pressure when facing extreme environmental situations. Fish from the Austrolebias group exhibit rapid neurogenesis in different brain regions, possibly as a result of the demanding conditions of a changing habitat. Knowledge of cerebral histology is essential for detecting ontogenic, anatomical, or cytoarchitectonic changes in the brain during the short lifespan of these fish, such as those reflecting functional adaptive plasticity in different systems, including sensory structures. The generation of an atlas of Garcialebias charrua (previously known as Austrolebias charrua) establishes its anatomical basis as a representative of a large group of fish that share similarities in their way of life. In this work, we present a detailed study of both gross anatomy and microscopic anatomy obtained through serial sections stained with the Nissl technique in three orientations: transverse, horizontal, and parasagittal planes. This atlas includes accurate drawings of the entire adult brain of the male fish Garcialebias charrua, showing dorsal, ventral, and lateral views, including where emergence and origin of cranial nerves. This brain atlas allows us to understand histoarchitecture as well as the location of neural structures that change during adult neurogenesis, enabling comparisons within the genus. Simultaneously, this atlas constitutes a valuable tool for comparing the brains of other fish species with different behaviors and neuroecologies.

Morphological evidence of the protective effects of a synthetic chalcone against the striatal myelin damage induced by glutaric acid.

Ultrastructural features of striatal white matter and cells in an in vivo model of glutaric acidemia type I created by intracerebral injection of glutaric acid (GA) were analyzed by transmission electron microscopy and immunohistochemistry. To test if the white matter damage observed in this model could be prevented, we administered the synthetic chemopreventive molecule CH38 ((E)-3-(4-methylthiophenyl)-1-phenyl-2-propen-1-one) to newborn rats, previous to an intracerebroventricular injection of GA. The study was done when striatal myelination was incipient and when it was already established (at 12 and 45 days post-injection [DPI], respectively). Results obtained indicate that that the ultrastructure of astrocytes and neurons did not appear significantly affected by the GA bolus. Instead, in oligodendrocytes, the most prominent GA-dependent injury defects included endoplasmic reticulum (ER) stress and nuclear envelope swelling at 12 DPI. Altered and reduced immunoreactivities against heavy neurofilament (NF), proteolipid protein (PLP), and myelin-associated glycoprotein (MAG) together with axonal bundle fragmentation and decreased myelin were also found at both ages analyzed. CH38 by itself did not affect striatal cells or axonal packages. However, the group of rats that received CH38 before GA did not show evidence neither of ER stress nor nuclear envelope dilation in oligodendrocytes, and axonal bundles appeared less fragmented. In this group, labeling of NF and PLP was similar to the controls. These results suggest that the CH38 molecule is a candidate drug to prevent or decrease the neural damage elicited by a pathological increase of GA in the brain. Optimization of the treatments and identification of the mechanisms underlying CH38 protective effects will open new therapeutic windows to protect myelin, which is a vulnerable target of numerous nervous system diseases.

Study of Trypanosoma cruzi epimastigote cell death by NMR-visible mobile lipid analysis.

Cell death mechanisms in Trypanosoma cruzi have not been disclosed in detail though different conventional techniques have been used in the classification of parasite-cell death type. Nuclear magnetic resonance (NMR) has successfully been used as a tool to evaluate the onset of apoptosis in a number of higher eukaryote-cell models analysing the ratio of CH(2)/CH(3) integration from the visible mobile lipids (VML). Surprisingly, this versatile non-invasive spectroscopy technique has never been employed with this purpose in T. cruzi. In the present study it is shown that under different parasite death-conditions the ratio CH(2)/CH(3) varied drastically. Thus, T. cruzi epimastigotes in apoptotic conditions increase significantly this ratio while in necrotic as well as in autophagic situations the parasites maintain the VML, CH(2)/CH(3) ratio, in normal values. Additionally, other VML markers commonly used in these studies, such as the change in the region of methyl-choline moiety, -N(+)(CH(3))(3), exhibited different particular patterns according to the type of cell death. Our results suggest that the (1)H NMR-VML technique is an adequate tool to discriminate different T. cruzi death pathways.

Plasticity of cell proliferation in the retina of Austrolebias charrua fish under light and darkness conditions.

Austrolebias annual fishes exhibit cell proliferation and neurogenesis throughout life. They withstand extreme environmental changes as their habitat dries out, pressuring nervous system to adapt. Their visual system is challenged to adjust as the water becomes turbid. Therefore, this study focused on how change in photic environment can lead to an increased cell proliferation in the retina. We administered 5-chloro-2'- deoxyuridine (CldU) and 5-iodo-2'-deoxyuridine (IdU) at different temporal windows to detect cell proliferation in natural light and permanent darkness. Stem/progenitor cells were recognized as IdU+/CldU + nuclei co-labeled with Sox2, Pax6 or BLBP found in the ciliary marginal zone (CMZ). The expression pattern of BLBP + glial cells and ultrastructural analysis indicates that CMZ has different cell progenitors. In darkness, the number of dividing cells significantly increased, compared to light conditions. Surprisingly, CMZ IdU+/CldU + cell number was similar under light and darkness, suggesting a stable pool of stem/progenitor cells possibly responsible for retinal growth. Therefore, darkness stimulated cell progenitors outside the CMZ, where Müller glia play a crucial role to generate rod precursors and other cell types that might integrate rod-dependent circuits to allow darkness adaptation. Thus, the Austrolebias fish retina shows great plasticity, with cell proliferation rates significantly higher than that of brain visual areas.

IMXQB-80: A Quillaja brasiliensis saponin-based nanoadjuvant enhances Zika virus specific immune responses in mice.

Vaccine adjuvants are compounds that enhance/prolong the immune response to a co-administered antigen. Saponins have been widely used as adjuvants for many years in several vaccines - especially for intracellular pathogens - including the recent and somewhat revolutionary malaria and shingles vaccines. In view of the immunoadjuvant potential of Q. brasiliensis saponins, the present study aimed to characterize the QB-80 saponin-rich fraction and a nanoadjuvant prepared with QB-80 and lipids (IMXQB-80). In addition, the performance of such adjuvants was examined in experimental inactivated vaccines against Zika virus (ZIKV). Analysis of QB-80 by DI-ESI-ToF by negative ion electrospray revealed over 29 saponins that could be assigned to known structures existing in their congener Q. saponaria, including the well-studied QS-21 and QS-7. The QB-80 saponins were a micrOTOF able to self-assembly with lipids in ISCOM-like nanoparticles with diameters of approximately 43 nm, here named IMXQB-80. Toxicity assays revealed that QB-80 saponins did present some haemolytical and cytotoxic potentials; however, these were abrogated in IMXQB-80 nanoparticles. Regarding the adjuvant activity, QB-80 and IMXQB-80 significantly enhanced serum levels of anti-Zika virus IgG and subtypes (IgG1, IgG2b, IgG2c) as well as neutralized antibodies when compared to an unadjuvanted vaccine. Furthermore, the nanoadjuvant IMXQB-80 was as effective as QB-80 in stimulating immune responses, yet requiring fourfold less saponins to induce the equivalent stimuli, and with less toxicity. These findings reveal that the saponin fraction QB-80, and particularly the IMXQB-80 nanoadjuvant, are safe and capable of potentializing immune responses when used as adjuvants in experimental ZIKV vaccines.

Synaptic vesicles in motor synapses change size and distribution during the day.

The morphology of Drosophila motor terminals changes along the day with a circadian rhythm controlled by the biological clock. Here, we used electron microscopy to investigate the size, number, and distribution of synaptic vesicles, at intervals of 6 h during 2 consecutive days, under light-dark (LD) or the first 2 days in constant darkness (DD). We found changes in the size and distribution of vesicles located either at the active zone or in the reserve pool, indicating a circadian rhythm of synapse reorganization. Vesicles at the active zone were generally smaller than those in the reserve pool in LD and DD conditions. The size of active zones vesicles decreased twice in LD, corresponding with times of more intense locomotion activity, but only once in DD conditions.

Value of quantitative ultramorphological sperm analysis in infertile men.

A specific cause of infertility cannot be identified in at least 25% of men referred to a specialized clinic. Diagnosis of infertile men is based mainly on standard semen analysis and the observation of sperm under light microscope. The aim of our study was to find the subcellular sperm characteristics that could explain infertility in a group of teratozoospermic infertile men. Morphological characteristics of sperm from non-teratozoospermic (control donors) and teratozoospermic infertile men were analyzed by transmission electron microscopy (TEM) and quantified. Our analysis showed that sperm cells from control donors presented a higher number of normal heads and tails than infertile men. Regarding subcellular characteristics of nucleus and tails, only the percentage of vacuolated nucleus, the absence of at least one pair of microtubules of the axoneme and the total distortion of the tail were statistically higher in infertile men than in control donors. There were no differences in the number of normal acrosomes between the groups. Although the ultrastructural sperm defects overlapped between control donors and infertile men, TEM permits the identification and differentiation of a larger amount of defects than light microscopy. Vacuolated nucleus and gross alterations of the tail are the major sperm defects that seem to have prognostic value in teratozoospermic men.

The Drosophila Nora virus is an enteric virus, transmitted via feces.

The biology of the Drosophila viruses has not been intensely investigated. Here we have investigated the biology of the Nora virus, a persistent Drosophila virus. We find that injected Nora virus is able to replicate in the files, reaching a high titer that is maintained in the next generation. There is a remarkable variation in the viral loads of individual flies in persistently infected stocks; the titers can differ by three orders of magnitude. The Nora virus is mainly found in the intestine of infected flies, and the histology of these infected intestines show increased vacuolization. The virus is excreted in the feces and is horizontally transmitted. The Nora virus infection has a very mild effect on the longevity of the flies, and no significant effect on the number of eggs laid and the percent of eggs that develop to adults.

Astrocytic proliferation and mitochondrial dysfunction induced by accumulated glutaric acidemia I (GAI) metabolites: possible implications for GAI pathogenesis.

Glutaric (GA) and 3-hydroxyglutaric (OHGA) acids accumulate in glutaric acidemia I (GAI), a neurometabolic disease characterized by acute striatal degeneration and chronic progressive cortical atrophy. To explore the hypothesis that astrocytes are involved in GAI pathogenesis and targets of accumulating metabolites, we determined the effects of GA and OHGA on cultured rat cortical astrocytes. Remarkably, both acids induced mitochondria depolarization and stimulated proliferation in confluent cultures without apparent cell toxicity. Newborn rats injected with GA systemically also showed increased cell proliferation in different brain regions. Most of the proliferating cells displayed markers of immature astrocytes. Antioxidant iron porphyrins prevented both mitochondria dysfunction and increased in vitro and in vivo proliferation, suggesting a role of oxidative stress in inducing astrocytosis. Taken together, the data suggest that mitochondrial dysfunction induced by GA metabolites causes astrocytes to adopt a proliferative phenotype, which may underlie neuronal loss, white matter abnormalities and macrocephalia characteristics of GAI.

Reduction of ischemic brain damage and increase of glutathione by a liposomal preparation of quercetin in permanent focal ischemia in rats.

Oxidative stress is implicated in the pathogenesis of cerebral ischemia injury, and the flavonoids have shown to be neuroprotective in experimental models of cerebral ischemia. Previously, we have shown that an aqueous preparation of quercetin did not reach the brain while a liposomal preparation produced measurable cerebral amounts of quercetin that reduced significantly the cerebral damage provoked by permanent middle cerebral artery occlusion (pMCAo) of rats. In this context, the protective effects of liposomal quercetin (LQ) were investigated in the same model after 1 and 4 hours of arterial occlusion. LQ was administered in a single dose (30 mg/kg), at 30 min, 1 and 4 h after pMCAo, and the brain was studied 24 h later. Cerebral damage and the oedema volume were assessed with a tetrazolium salt (TTC). The status of brain tissue, the neuronal population, the global motor behaviour as well as the antioxidant, endogenous reduced glutathione (GSH), were also assessed in the brain. Thirty min after LQ there was a significantly protective effect against ischemic lesion demonstrated by a significant increase in numbers of cells in striatum and cortex, together with a partial reversal of motor deficits. GSH levels decreased after ischemia in ipsilateral striatum and cortex, and the LQ preparation reversed these effects 24 h after the occlusion. Our results suggest that endogenous brain GSH is critical in the defense mechanisms after ischemia, as a significant mediator of the protective effects of the LQ preparation.

Kinesin 1 regulates cilia length through an interaction with the Bardet-Biedl syndrome related protein CCDC28B.

Bardet-Biedl syndrome (BBS) is a ciliopathy characterized by retinal degeneration, obesity, polydactyly, renal disease and mental retardation. CCDC28B is a BBS-associated protein that we have previously shown plays a role in cilia length regulation whereby its depletion results in shortened cilia both in cells and Danio rerio (zebrafish). At least part of that role is achieved by its interaction with the mTORC2 component SIN1, but the mechanistic details of this interaction and/or additional functions that CCDC28B might play in the context of cilia remain poorly understood. Here we uncover a novel interaction between CCDC28B and the kinesin 1 molecular motor that is relevant to cilia. CCDC28B interacts with kinesin light chain 1 (KLC1) and the heavy chain KIF5B. Notably, depletion of these kinesin 1 components results in abnormally elongated cilia. Furthermore, through genetic interaction studies we demonstrate that kinesin 1 regulates ciliogenesis through CCDC28B. We show that kinesin 1 regulates the subcellular distribution of CCDC28B, unexpectedly, inhibiting its nuclear accumulation, and a ccdc28b mutant missing a nuclear localization motif fails to rescue the phenotype in zebrafish morphant embryos. Therefore, we uncover a previously unknown role of kinesin 1 in cilia length regulation that relies on the BBS related protein CCDC28B.

Neuromuscular junction in abdominal muscles of Drosophila melanogaster during adulthood and aging.

The neuromuscular junction (NMJ) of Drosophila melanogaster has been established as a productive model for the study of synaptogenesis, synaptic plasticity, vesicle recycling, and other synaptic functions in embryos and larvae. It also has potential for the study of long-term plasticity during adult life and degenerative processes associated with aging. Here we provide a detailed description of the morphology and ultrastructure of the NMJ on abdominal dorsal longitudinal muscles throughout adult life from eclosion to senescence. In contrast to the case in the larva, the predominant type of terminals in these muscles in the adult fly consists of only two or three branches with tightly packed synaptic boutons. We observed qualitative and quantitative changes as mean bouton size increased gradually during adulthood, and the largest boutons were present in the old fly. The length of nerve branches first increased and thereafter decreased gradually during most of adult life. Branch diameter also decreased progressively, but branch number did not change. The subsynaptic reticulum became progressively thinner, and "naked" boutons were found in old flies. Ultrastructural traits gave indications of an age-associated increment in autophagy, larger synaptic vesicles, and impaired endocytosis. We propose that NMJ aging in the fly correlates with impaired endocytosis and membrane dynamics. This view finds a functional correlate in flies carrying a temperature-sensitive mutation in shibire that reversible blocks endocytosis; age significantly reduces the time required for complete paralysis and increases the time of recovery, thus confirming the age-dependent alteration in vesicle dynamics.

Stem cells distribution, cellular proliferation and migration in the adult Austrolebias charrua brain.

Our previous studies demonstrated that Austrolebias charrua annual fish is an excellent model to study adult brain cell proliferation and neurogenesis due to the presence of active and fast neurogenesis in several regions during its short lifespan. Our main goal was to identify and localize the cells that compose the neurogenic areas throughout the Austrolebias brain. To do this, we used two thymidine halogenated analogs to detect cell proliferation at different survival times: 5-chloro-2'-deoxyuridine (CldU) at 1day and 5-iodo-2'-deoxyuridine (IdU) at 30days. Three types of proliferating cells were identified: I - transient amplifying or fast cycling cells that uptake CldU; II - stem cells or slow cycling cells, that were labeled with both CldU and IdU and did not migrate; and III - migrant cells that uptake IdU. Mapping and 3D-reconstruction of labeled nuclei showed that type I and type II cells were preferentially found close to ventricle walls. Type III cells appeared widespread and migrating in tangential and radial routes. Use of proliferation markers together with Vimentin or Nestin evidenced that type II cells are the putative stem cells that are located at the ventricular lumen. Double label cells with IdU+ and NeuN or HuC/D allowed us identify migrant neurons. Quantitation of labeled nuclei indicates that the proportion of putative stem cells is around 10% in all regions of the brain. This percentage of stem cells suggests the existence of a constant brain cell population in Austrolebias charrua that seems functional to the maintainance of adult neurogenesis.

Characterization of primary cilia during the differentiation of retinal ganglion cells in the zebrafish.

BACKGROUND: Retinal ganglion cell (RGC) differentiation in vivo is a highly stereotyped process, likely resulting from the interaction of cell type-specific transcription factors and tissue-derived signaling factors. The primary cilium, as a signaling hub in the cell, may have a role during this process but its presence and localization during RGC generation, and its contribution to the process of cell differentiation, have not been previously assessed in vivo. METHODS: In this work we analyzed the distribution of primary cilia in vivo using laser scanning confocal microscopy, as well as their main ultrastructural features by transmission electron microscopy, in the early stages of retinal histogenesis in the zebrafish, around the time of RGC generation and initial differentiation. In addition, we knocked-down ift88 and elipsa, two genes with an essential role in cilia generation and maintenance, a treatment that caused a general reduction in organelle size. The effect on retinal development and RGC differentiation was assessed by confocal microscopy of transgenic or immunolabeled embryos. RESULTS: Our results show that retinal neuroepithelial cells have an apically-localized primary cilium usually protruding from the apical membrane. We also found a small proportion of sub-apical cilia, before and during the neurogenic period. This organelle was also present in an apical position in neuroblasts during apical process retraction and dendritogenesis, although between these stages cilia appeared highly dynamic regarding both presence and position. Disruption of cilia caused a decrease in the proliferation of retinal progenitors and a reduction of neural retina volume. In addition, retinal histogenesis was globally delayed albeit RGC layer formation was preferentially reduced with respect to the amacrine and photoreceptor cell layers. CONCLUSIONS: These results indicate that primary cilia exhibit a highly dynamic behavior during early retinal differentiation, and that they are required for the proliferation and survival of retinal progenitors, as well as for neuronal generation, particularly of RGCs.

Telencephalic-olfactory bulb ventricle wall organization in Austrolebias charrua: Cytoarchitecture, proliferation dynamics, neurogenesis and migration.

Adult neurogenesis participates in fish olfaction sensitivity in response to environmental challenges. Therefore, we investigated if several populations of stem/progenitor cells that are retained in the olfactory bulbs (OB) may constitute different neurogenic niches that support growth and functional demands. By electron microscopy and combination cell proliferation and lineage markers, we found that the telencephalic ventricle wall (VW) at OB level of Austrolebias charrua fish presents three neurogenic niches (transitional 1, medial 2 and ventral 3). The main cellular types described in other vertebrate neurogenic niches were identified (transient amplifying cells, stem cells and migrating neuroblasts). However, elongated vimentin/BLBP+ radial glia were the predominant cells in transitional and ventral zones. Use of halogenated thymidine analogs chloro- and iodo-deoxyuridine administered at different experimental times showed that both regions have the highest cell proliferation and migration rates. Zone 1 migration was toward the OB and telencephalon, whereas in zone 3, migration was directed toward the OB rostral portion constituting the equivalent of the mammal rostral migratory band. Medial zone (MZ) has fewer proliferating non-migrant cells that are the putative stem cells as indicated by short and long proliferation assays as well as cell lineage markers. Sparse migration observed suggests MZ may collaborate with VW growth. Scanning electron microscopy evidenced that the whole VW has only monociliated cells with remarkable differences in cilium length among regions. In OB there are monociliated cells with dwarf cilium whereas ventral telencephalon shows long cilium. Summarizing, we identified three neurogenic niches that might serve different functional purposes.

Morphological evaluation of sperm from infertile men selected by magnetic activated cell sorting (MACS).

Electron microscopy analysis performed in five infertile human subjects after sperm selection by swim-up followed by magnetic activated cell sorting (MACS) demonstrated a decrease in the number of spermatozoa with characteristics compatible with cell death. However, no significant differences were found when the swim-up/MACS semen fraction was compared with swim-up fraction alone.