Stem cells and neuronal repair

Bacterial infection of the brain: how bacteria penetrate the CNS by invading peripheral nerves

Objective: To investigate the effects of neonatal stroke on progenitor cells lining the lateral ventricles. Methods: Intraventricular injection of replication-incompetent green fluorescent protein (GFP)-expressing lentivirus was performed in postnatal day 1 (P1) rats to specifically label radial glia/type B neural stem cells and ependymal cells of the lateral ventricle. A subset of animals was exposed to transient middle cerebral artery occlusion (MCAO) at P7, with mild or moderate injury confirmed by diffusion-weighted MRI and histology. Newborn cells were identified by GFP expression, location and expression of cell type-specific markers in the striatum, cortex and olfactory bulb using confocal microscopy and systematic random sampling. Results: Three weeks lentiviral GFP transduction of cells in the lateral ventricle, abundant GFP-expressing neurons and glia were identified in the rostral migratory stream, olfactory bulb and striatum as expected from labeling the subventricular zone (SVZ) type B neural stem cell lineage. Two weeks following mild or severe focal stroke at P7, no GFP-expressing neurons were detected in striatum or cortex although some single-labeled doublecortin+ immature neurons were detected in the penumbra. The densities of GFP+/ glial fibrillary acidic protein (GFAP)+ astrocytes and GFP+/O4+ oligodendrocytes were reduced in the striatum following MCAO (4.8 ± 1.02 vs. 2.5 ± 0.4 cells/high-power field, HPF; p = 0.005; 2.8± 1 vs. 0.5 ± 0.2 cells/HPF, p = 0.008). Furthermore, there was a reduction of GFP+ cells in the olfactory bulb following MCAO (58.8 ± 14.9 vs. 19.6 ± 5.4 cells/HPF, p = 0.025). Finally, there was an increased percentage of GFP+/GFAP+ cells (70 vs. 50%), with a decreased proportion of GFP+/O4+ cells (14 vs. 30%) in injured animals. Conclusion: Neurogenesis originating from cells of the lateral ventricle, including SVZ type B cells, is significantly reduced following neonatal stroke. Furthermore, neonatal stroke disrupts gliogenesis in the striatum, decreasing overall numbers of new glia and shifting the population towards astrocytes.

The extrinsic and intrinsic factors involved in regulating the developmental progression from totipotent embryonic stem cells to phenotypically restricted neural stem and progenitor cells have been subjects of intense research over the last decade. Two major reasons for this concentrated effort are the gradual acceptance that at least some regions of the brain generate neurons throughout life, and the growing appreciation that these cells could be used to therapeutically treat disorders and injuries of the central nervous system (CNS). Despite early studies by Altman and Das (1) demonstrating ongoing neurogenesis in the adult rodent hippocampus and olfactory bulb, it was widely believed until recently that in mammals the generation of neurons ceases in the late embryonic or early postnatal period. In addition to the persistent neurogenesis in the olfactory bulb and hippocampus, olfactory receptor neurons (ORNs), the first-order neurons in the peripheral olfactory system, were also found to regenerate throughout life in all vertebrates examined (2–4). Postnatal neurogenesis also occurs in the neonatal cerebellum (see ref. 1), although it is more limited in duration than that of the olfactory bulb, the hippocampus, and the ORNs. Populations of neural progenitor cells, which generate the neurons and/or glia in these regions with extended proliferation, have now been identified in the postnatal brain (5–11).

Objective To investigate the possible abnormal changes of adult neurogenesis in the rostral migratory stream(RMS) , in the olfactory bulb and in learning or memory of renovascular hypertensive rat, and to evaluate the possible efficacies produced by intervention of exogenous melatonin. Methods Renovascular hypertensive rat model was established by clamping bilateral renal arteries. Rats were randomly divided into four groups with 10 rats per group: hypertensive, sham-operated, melatonin-treated and normal groups. Morris water maze was used to detect the abilities of learning and memory, and immunohistochemistry for bromodeoxyuridine(BrdU), bromodeoxyuridine plus glial fibrillary acidic protein (BrdU+ GFAP) and bromodeoxyuridine plus neurofilament (BrdU+ NT) was utilized to examine the changes of the adult neurogenesis in RMS and in the olfactory bulb of each rat. Results The escape latency of hypertensive group ((29.95±20.11) s) recorded in Morris water maze test was the longest in a significant comparison to those of normal group((20.58±19.18)s), sham-operated group ((22.11±20.28)s) and melatonin-treated group((23.81±22.25)s)(P<0.05); while the time spent by rats during swimming in the quadrant with being originally put platform of hypertensive((25.51±5.21)s) was the shortest by significant comparison to those of normal((34.67±4.53)s), sham-operated((33.56±6.12)s) and melatonin-treated group ((36.23±3.86)s)(P<0.05). Proliferation of neural stem cells(cells/field): the number of BrdU immunoreactive (BrdU-IR) cells of rats in RMS of hypertensive group(34.08±6.49) was the smallest, significantly compared with those of normal(43.53±7.56), sham-operated(46.11±4.75) and melatonin-treated group(44.30±8.29), (P<0.05). Differentiation of neural stem cells(cells/slice): the number of BrdU+ GFAP-IR cells in olfactory bulb of hypertensive group(2.45±1.32) was largest, significantly compared with those of normal(1.01±0.78), sham-operated(0.68±0.37) and melatonin-treated group(1.35±0.88)(P<0.05); while the number of BrdU+ NF-IR cells in olfactory bulb of hypertensive group(1.15±0.81) was the smallest, significantly compared with those of normal (1.89±0.98), sham-operated(1.63±1.01) and melatonin-treated group(2.30±1.22)(P<0.05). Conclusion Melatonin can ameliorate the abnormal cognitive behaviors induced by hypertension via a mechanism of reversing the progression of neurogenesis disorders both in RMS and in olfactory bulb, indicating that melatonin may prevent the development of dementia triggered by hypertension. Key words: Melatonin; Hypertension; Spatial memory; Rostral migratory stream; Olfactory bulb

Most organisms rely on an olfactory system to detect and analyze chemical cues from the external world in the context of essential behavior. From worms to vertebrates, chemicals are detected by odorant receptors expressed by olfactory sensory neurons, which send an axon to the primary processing center—the olfactory bulb, in vertebrates. Within this relay, sensory neurons form excitatory synapses with projection neurons and with inhibitory interneurons. Thus, due to complex synaptic interactions in the olfactory bulb circuit, the output of a given projection neuron is determined not only by the sensory input, but also by the activity of local inhibitory interneurons that are concerned by adult neurogenesis throughout life. Recent studies have provided clues about how these new neurons incorporate into preexisting networks, how they survive or die once integrated into proper microcircuits, and how basic network functions are maintained despite the continual renewal of a large percentage of neurons. We know that external influences modulate the process of late neurogenesis at various stages. Thus, this process is probably flexible, allowing brain performance to be optimized for its environment. But optimized how? And why? This chapter describes the adaptation of new interneuron production to experience-induced plasticity. In particular, how the survival of newly generated neurons is highly sensitive not only to the level of sensory inputs, but also to the behavioral context is discussed. Also discussed is how neurogenesis may finely tune the functioning of the neural network, optimizing the processing of sensory information. Adult neurogenesis maintains continual turnover...

It had long been held that neurogenesis in the mammalian central nervous system (CNS) was largely completed by birth. This opinion held sway until the early 1960s, when cellular proliferation was discovered in the subventricular zone (SVZ) (1) and neurogenesis was observed to occur in the olfactory bulb (2,3). The significance of these studies went unexplored until recently, when the ability to extract, grow, and reimplant neural stem cells (NSCs) into the brain forced a re-examination of the intrinsic capacity of the cerebrum to marshal endogenous stem cell pools (4–6). Similarly for the spinal cord, Adrian and Walker (7), employing 3H-thymidine, a mitotic marker, labeled a small population of cells lining the central canal of the intact adult rat. Because these cells had only a limited life-span, however, their role was deemed to be trivial. The significance of this observation for spinal cord dysfunction and repair has also recently been revisited in light of the possible existence of NSCs within the spinal cord.

Objective To investigate the effects of electrical stimulation of olfactory bulb (OB)on proliferation,migration to OB of neural precursor cells (NPC) in the subventricular zone,and preliminary explore its related mechanism.Methods Eighty adult female Sprague Dawley rats were randomly divided into normal control group,sham stimulation group and stimulation groups of 1 d,3 d,1 week,2 weeks,3 weeks and 4 weeks (n=10).Rats in the later six groups were performed OB electrical stimulation.Five of these rats in each group were injected intraperitoneally with 5-bromo-2-deoxyuridine (Brdu) to mark new-bom cells and immunohistochemical staining was employed to observe the proliferation of NPCs in SVZ; the other 5 were sacrificed at corresponding time to detect the mRNA expression ofprokineticin 2 (prk2) in OB.Another 15 rats were randomly divided into normal control group Ⅰ,sham stimulation group Ⅰ and stimulation group Ⅰ (n=5); four weeks after Brdu was injected,the animals were sacrificed and immumohistochemical staining was used to investigate the number of Brdu-positive cells in OB.Results The number of Brdu-positive cells in SVZ was significantly different among the eight group (F=51.475,P=0.000); as compared with those in the normal control group and sham stimulation group,the Brdu-positive cells were significantly increased in stimulation groups of 1 d,3 d,1 week and 2 weeks (P＜0.05).The mRNA expression ofprk 2 in OB was significantly different among the eight group (F=154.067,P=0.000); as compared with those in the normal control group and sham stimulation group,the Brdu-positive cells were significantly increased in all the stimulation groups (F=36.472,P=0.000).Four weeks after injection of Brdu,the Brdu-positive cells in OB of the stimulation group Ⅰ significantly increased as compared with that in the normal control group Ⅰ and sham stimulation group Ⅰ (P＜0.05).Conclusion Electrical stimulation of OB promotes proliferation,migration of NPCs in SVZ,which may be related to elevated expression of Prk2. Key words: Electrical stimulation; Olfactory bulb; Neural precursor cell; Prokineticin 2

Over most of the past century, it was thought that the adult brain was completely incapable of generating new neurons. However, in the last decade, the development of new techniques has resulted in an explosion of new research showing that (i) neurogenesis, the birth of new neurons, is not restricted to embryonic development, but normally also occurs in two limited regions of the adult mammalian brain (the olfactory bulb and the dentate gyrus of the hippocampus); (ii) that there are significant numbers of multipotent neural precursors in many parts of the adult mammalian brain; and (iii) that it is possible to induce neurogenesis even in regions of the adult mammalian brain, like the neocortex, where it does not normally occur, via manipulation of endogenous multipotent precursors in situ. In the neocortex, recruitment of small numbers of new neurons can be induced in a region-specific, layer-specific, and neuronal type-specific manner, and newly recruited neurons can form long-distance connections to appropriate targets. This suggests that elucidation of the relevant molecular controls over adult neurogenesis from endogenous neural precursors/stem cells may allow the development of neuronal replacement therapies for neurodegenerative disease and other central nervous system injuries that may not require transplantation of exogenous cells.

Objective To investigate the effects of electrical stimulation of olfactory bulb( OB) on the proliferation, migration and differentiation of neural stem cell ( NSC ) in subventricular zone. Method Forty - eight adult female Sprague - Dawley(SD) rats were randomly divided into control group, sham stimulation group and stimulation group including 6 time points(1 day,3 day,7 day, 14 day,21 day, 28 day). The rats were injected intraperitoneally with bromodeoxyuridine( BrdU) to detect the proliferation of NSC by immunohistochemistry staining. Another 18 rats were randomly divided into control group, sham stimulation group and stimulation group. Four weeks after BrdU injection, the rats were sacrificed and immunohistochemistry and immunofluorescence were used to investigate the migration and differentiation of NSC in OB. Results In SVZ, BrdU - positive cells began to increase 1 d after stimulation (17. 67 ± 1.03, P <0.01) .reached to the maximum level at 1 week(28. 50 ± 1. 87, P <0. 01) ,then decreased to normal at 3 week. Four weeks after injection of BrdU,the BrdU -positive cells significantly increased in RMS(67. 33 ±3.50, P <0.01) and OB(44.33 ±5.47, P <0.01) in stimulation group. Fluorescence double staining showed that the stimulation of OB had no effect on the differentiation of NSC into neurons or gliocytes. Conclusions Electrical stimulation of OB could promote proliferation, migration of NSC in SVZ, but it has no effect on the differentiation of NSC into neurons or gliocytes. Key words: Electrical stimulation; Olfactory bulb; Neurogenesis; Rats

To analyze the correlation between olfactory bulb (OB) volumes and ages, as well as OB volumes and olfactory function in healthy adults. One hundred healthy subjects, without any diseases which might cause smell dysfunction were selected to participate in this study. The ages of the subjects ranged from 20 to 70, with the mean age of 42.6 ± 4.8. These subjects were scaned for olfactory function by T&T testing, OB volumes assessed with Magnetic resonance imaging (MRI). The correlation between OB volumes and ages as well as OB volumes and olfactory function in healthy adults was analyzed. SPSS 13.0 software was used to analyze the data. The left and right OB volumes of men were (84.65 ± 7.11) mm(3) and (87.79 ± 7.57) mm(3), average OB volume was (86.14 ± 7.37) mm(3). The left and right OB volumes of women were (69.58 ± 4.72) mm(3) and (71.43 ± 5.29) mm(3), average OB volume was (70.22 ± 5.02) mm(3). OB volume study revealed no statistical difference between the left and right OB volumes of men (t = 1.024, P > 0.05). OB volume study revealed no statistical difference between the left and right OB volumes of women (t = 0.987, P > 0.05). OB volumes were lower in women as compared with men (t value were 3.742, 3.869 and 3.814, all P < 0.01). OB volumes were negatively correlated with ages in the subjects (r value were -0.588, -0.523, both P < 0.01). Olfactory discriminate threshold was negatively correlated with OB volumes in the subjects (r value were -0.624, -0.587, both P < 0.01). This study revealed no statistical difference between the left and right OB volumes in healthy adults. OB volumes were lower in women as compared with men. As age went up, OB volume would become smaller. As olfactory function went down, OB volume would become smaller. OB volumes were expected to be a gauge of olfactory function.

The tight junction protein claudin-3 is overexpressed in diverse epithelial tumours and is associated with increased survival, progression and motility of tumour cells. Claudin-3 expression profiles are being increasingly used for diagnostic and prognostic tumour classification. Claudin-3 has been identified as a receptor for Clostridium perfringens enterotoxin, which is under consideration for selective lysis of claudin-3-expressing tumours, particularly brain metastases, and other translational medicine uses. However, the localization of claudin-3 in the brain has not been completely elucidated. While claudin-3 in brain tissue adjacent to claudin-3-expressing metastases had been excluded and low or undetectable levels proposed in the CNS, under physiological conditions, in adult human, rat and mouse brains, claudin-3 was exclusively found in choroid plexus epithelium where it is considered an integral component of the blood-cerebrospinal-fluid barrier. We report here the pronounced presence of claudin-3 not only in the nasal region (as described for rat), but also in the mouse olfactory bulb and nerve using immunohistochemistry and Western blot. Claudin-3 was present in the fila olfactoria from the epithelium to the olfactory nerve and in the main and accessory olfactory bulb. We propose that the abundant presence of claudin-3 in the olfactory system, particularly in nerve fibres and the olfactory bulb cone, which we present here, may play a role at the interface of the central and peripheral nervous system, both as barrier and for axonal growth and communication. Thus, claudin-3 should be considered and further explored with regards to treatment approaches addressing the olfactory bulb and nasal region.

Activity-dependent synaptic structural plasticity underlies the learning and memory. Mammals, especially the rodents, are very sensitive to odorants, and have considerable capability of odor learning and memory. Here, the activity-dependent synaptic structural plasticity in the olfactory bulb (OB) of the CNGA2 knock-out transgenic mice (CNGA2 KO), which is anosmic, was investigated. Using immunohistochemistry for specific presynaptic and postsynaptic markers, it was found that deficits of peripheral inputs induced significant decreases in the expression of synaptophysin, a general marker for synapses, and gephyrin, a marker for inhibitory synapses, in the external plexiform layer (EPL) and the granule cell layer (GCL), but the vesicular glutamate transporters 1 (VGluT1) decreased only in EPL, not in GCL. Western-blots showed the decreases in the expression of gephyrin in the OB of CNGA2 KO mice, but not in the expression of the VGluT1. The results of immunohistochemistry and Western blot revealed that the excitatory and inhibitory synapses may have changed after deficits of peripheral inputs. GCs were the main participants in the EPL and GCL in the OB. Dendritic spines are the postsynaptic sites of the majority of excitatory synapses in the mammalian central nervous system, and the morphology and dynamics of dendritic spines change in response to novel experiences and neuropathologies. In the OB, spines on mature GCs are recipients of glutamatergic synapses in the GCL and reciprocal synapses in the EPL. Almost all study related to structural plasticity of GCs concentrated on the adult-born GCs, but the number of new-born granule cells in the OB is negligible compared with the number of preexisting GCs. In order to further reveal the quantitive changes of glutamatergic synapses on GCs, in vivo adult brain plasmid electroporation to label mature GCs in the OB directly were adopted. Here, the spines in EPL were defined as distal spines and the spines in GCL as proximal spines. It was revealed that the density of spines on granule cells decreased significantly in EPL (distal spines) of CNGA2 KO mice, but did not change significantly in GCL (proximal spines), as same as the result of optical density analysis in the VGluT1 immunolabeling. These data suggest that the structural plasticity of the distal dendrodendritic synapses, rather than the proximate axon-dendritic synapses on granular cells of the OB, are significantly affected by the peripheral olfactory inputs.

In the adult mammalian brain, new neurons are added to the olfactory bulb (OB) throughout life. In rodents, the adult germinal region for OB neurogenesis is the subventricular zone (SVZ), a layer of cells found along the walls of the brain lateral ventricles (for review, see Alvarez-Buylla and Garcia-Verdugo 2002). Neuroblasts born in the SVZ migrate a relatively long distance into the OB where they then disperse radially and differentiate into interneurons. Most of these new OB neurons integrate into functional circuits (Belluzzi et al. 2003; Carleton et al. 2003), and about half survive long-term (Petreanu and Alvarez-Buylla 2002). SVZ cell proliferation is lifelong (Kuhn et al. 1996; Goldman et al. 1997; Molofsky et al. 2006), with thousands of new neurons generated daily for the mouse OB (Lois and Alvarez-Buylla 1994). The adult SVZ is also the birthplace of oligodendrocytes in both normal and diseased brain (Nait-Oumesmar et al. 1999; Picard-Riera et al. 2002; Menn et al. 2006; Parent et al. 2006). This profound level of continuous neurogenesis and concomitant oligodendrogliogenesis argues for the existence of a self-renewing multipotent precursor cell—or, neural stem cell (NSC)—within the SVZ. The SVZ-OB system is an attractive model in which to study neurogenesis and neuronal replacement as it includes the basic processes of NSC maintenance, progenitor cell-fate specification, migration, differentiation, and survival/death of newly born neurons. The enduring quality and stable cytoarchitecture of adult SVZ-OB neurogenesis may make these complex biological processes experimentally more tractable in comparison to studies of embryonic brain...

Over the last 50 yr, the long-held dogma that neurogenesis stops at birth has been gradually modified to allow specific exceptions to the rule. In many mammalian species, new neurons are continually added to the adult olfactory bulb (1) and hippocampus (2–5). As more information is gathered, additional sites may be found that are unique to individual species (6). Although natural neurogenic processes respond in a limited manner to damage in the CNS (7–9) the inability of the adult brain to regenerate fully following disease or injury still provides a concrete example of how the dogma holds true from a clinical point of view. In this context, the insight gained in defining neurogenic mechanisms in the adult may be relevant to central nervous system (CNS) repair since the successful therapeutic approaches will ultimately depend on the precise modulation of an extensive regulatory network.

Objective To demonstrate the morphology of drainage of the cerebrospinal fluid through the olfactory nerves into the cervical lymphatics. Methods Microfil was injected into the cisterna magna of the rabbit, cervical lymphatics, the olfactory bulbs, olfactory tracts and the nasal mucosa were observed by macroscopic anatomy and light microscopy. Results Under the operative microscope, Microfil was found within the subarachnoid space and along the olfactory nerves. At the nasal mucosa, a lymphatic network stained in yellow was identified near the olfactory nerves, which cross nasopharyngeal region and finally emptied into the superficial and deep cervical lymph nodes. Light microscopically, Microfil was distributed around the olfactory nerves and within the lymphatic vessels. At the nasal mucosa, the lymphatics were frequently located close to the nerves. Conclusion These results indicate that the cerebrospinal fluid drains from the subarachnoid space along the olfactory nerves to the nasal Lymphatics, which in turn, empties into the cervical lymph nodes. This anatomical communication, thus, allows the central nervous system to connect with the lymphatic system. The presence of this route may play an important role in immune disease of the central nervous system and regulation of the cerebrospinal fluid circulation. Key words: Lymph; Cerebrospinal fluid; Subarachnoid space; Central nervous system