Efficacy of Dequalinium Chloride vs Metronidazole for the Treatment of Bacterial Vaginosis: A Randomized Clinical Trial.

Importance: Bacterial vaginosis (BV) is a common cause of vaginal infection. First-line treatments of BV are metronidazole and clindamycin. Due to the increase in antibiotic resistance, effective nonantibiotic treatments for BV are needed. Objective: To examine whether dequalinium chloride, a broad-spectrum antiseptic, is noninferior to oral metronidazole for the treatment of BV. Design, Setting, and Participants: This phase 4, multicenter, triple-blind, double-dummy, parallel, noninferiority randomized clinical trial was conducted from July 29, 2021, to August 25, 2022, with a 1-month follow-up. Participants were premenopausal women 18 years or older with BV from 11 gynecologic practices and 1 hospital in Poland, Slovakia, and the Czech. Intervention: Patients were randomized to treatment with dequalinium chloride vaginal tablets (10 mg once daily for 6 days) or oral metronidazole (500 mg twice daily for 7 days). Double-dummy medication kits contained vaginal and oral tablets with placebo and active medication. Main Outcomes and Measures: The main outcome was the noninferiority margin (of 15 percentage points) in the absolute difference in clinical cure rates between dequalinium chloride and metronidazole 7 to 11 days after start of treatment (visit 1). Noninferiority was met if the lower 95% CI for the difference in clinical cure rate was less than 15 percentage points at visit 1. Results: A total of 147 women (mean [SD] age, 36.7 [9.0] years) were treated with dequalinium chloride (n = 72) or metronidazole (n = 75). The clinical cure rates at visit 1 were 64 of 69 (92.8%) for dequalinium chloride vs 69 of 74 (93.2%) for metronidazole in the intention-to-treat population, whereas in the per-protocol population, cure rates were 54 of 58 (93.1%) for dequalinium chloride vs 48 of 53 (90.6%) for metronidazole. The treatment differences of -0.5 percentage points (95% CI, -10.8 to 9.8 percentage points; P = .002) in the intention-to-treat population and 2.5 percentage points (95% CI, -9.4 to 14.4 percentage points; P = .001) in the per-protocol population confirmed the noninferiority of dequalinium chloride. The tolerability of dequalinium chloride was rated as very good by 30 of 50 patients (60.0%) but only by 21 of 54 (38.9%) for metronidazole. Three patients in the metronidazole group suspended treatment due to an adverse event. Conclusions and Relevance: This randomized clinical trial showed that dequalinium chloride was not inferior to metronidazole for the treatment of BV. Dequalinium chloride had a similarly high cure rate but with better tolerability and fewer adverse events. With a similar efficacy to metronidazole and clindamycin, dequalinium chloride warrants consideration as first-line treatment for BV to help reduce antibiotic consumption. Trial Registration: EudraCT: 2020-002489-15.

Neural stem cells in the adult olfactory bulb core generate mature neurons in vivo.

Although previous studies suggest that neural stem cells (NSCs) exist in the adult olfactory bulb (OB), their location, identity, and capacity to generate mature neurons in vivo has been little explored. Here, we injected enhanced green fluorescent protein (EGFP)-expressing retroviral particles into the OB core of adult mice to label dividing cells and to track the differentiation/maturation of any neurons they might generate. EGFP-labeled cells initially expressed adult NSC markers on days 1 to 3 postinjection (dpi), including Nestin, GLAST, Sox2, Prominin-1, and GFAP. EGFP+ -doublecortin (DCX) cells with a migratory morphology were also detected and their abundance increased over a 7-day period. Furthermore, EGFP-labeled cells progressively became NeuN+ neurons, they acquired neuronal morphologies, and they became immunoreactive for OB neuron subtype markers, the most abundant representing calretinin expressing interneurons. OB-NSCs also generated glial cells, suggesting they could be multipotent in vivo. Significantly, the newly generated neurons established and received synaptic contacts, and they expressed presynaptic proteins and the transcription factor pCREB. By contrast, when the retroviral particles were injected into the subventricular zone (SVZ), nearly all (98%) EGFP+ -cells were postmitotic when they reached the OB core, implying that the vast majority of proliferating cells present in the OB are not derived from the SVZ. Furthermore, we detected slowly dividing label-retaining cells in this region that could correspond to the population of resident NSCs. This is the first time NSCs located in the adult OB core have been shown to generate neurons that incorporate into OB circuits in vivo.

Segmental Mandibular Reconstruction Using Tissue Engineering Strategies: A Systematic Review of Individual Patient Data.

OBJECTIVE: The aim of the systematic review was to analyze the current clinical evidence concerning the use of tissue engineering as a treatment strategy for reconstruction of segmental defects of the mandible and their clinical outcomes using individual patient data. METHODS: A systematic review of the literature was conducted using PubMed and Cochrane Library on May 21, 2019. The eligibility criteria included patients in whom segmental mandibular reconstruction was carried out using tissue engineering as the primary treatment strategy. After screening and checking for eligibility, individual patient data were extracted to the extent it was available. Data extraction included the type of tissue engineering strategy, demographics, and indication for treatment, and outcomes included clinical and radiographic outcome measures, vitality of engineered bone, dental rehabilitation, and patient-reported outcome measures and complications. RESULTS: Out of a total of 408 articles identified, 44 articles reporting on 285 patients were included, of which 179 patients fulfilled the inclusion criteria. The different tissue engineering treatment strategies could be broadly classified into 5 groups: "prefabrication," "cell culture," "bone morphogenetic protein (BMP) without autografts," "BMP with autografts," and "scaffolds containing autografts." Most included studies were case reports or case series. A wide variety of components were used as scaffolds, cells, and biological substances. There was not a single outcome measure that was both objective and consistently reported, although most studies reported successful outcome. DISCUSSION: A wide variety of tissue engineering strategies were used for segmental mandibular reconstruction that could be classified into 5 groups. Due to the low number of treated patients, lack of standardized and consistent reporting outcomes, lack of comparative studies, and low evidence of reported literature, there is insufficient evidence to recommend any particular tissue engineering strategy.

Predictors of mortality after pelvic fractures: a retrospective cohort study from a level one trauma centre in Upper Egypt.

PURPOSE: The primary objective was to identify the predictors of in-hospital mortality after pelvic ring injuries. Secondary objectives were to analyze the differences between adults and children and to analyze the causes and timing of death. METHODS: A retrospective cohort study from the pelvic registry of Assiut University Trauma Unit (AUTU), a level 1 trauma centre in Upper Egypt, was carried out. A total of 1188 consecutive patients with pelvic ring fractures treated from January 2010 to December 2013 were eligible for analysis. Potential predictors were identified using standard statistical tests: univariable and multivariable regression analysis. RESULTS: Nine hundred fifty-one were adults (above 16 years) and 237 were children. According to Tile's classification, fractures type A, B, and C were 31.8%, 25.1%, and 43.1%, respectively. About a third of patients had fractures with soft tissue injury. Abdominopelvic collection as diagnosed by Focused Assessment with Sonography for Trauma (FAST) was positive in 11%. Associated injuries were present in 67.3% with abdominal-urogenital injuries being the most prevalent (66.3%). Median hospital stay was five days. Fifty-two patients (4.4%) were admitted to the ICU. One hundred three patients died (8.7%) within two peaks: first 24 hours and between 48 hours and one week. Multivariable logistic regression analysis identified increasing age, fractures with soft tissue injury, associated head injury, positive FAST examination, and admission to an ICU as significant predictors of in-hospital mortality. CONCLUSIONS: The first 24 hours were confirmed to be critical for survival in pelvic fracture patients. Advancing age, associated soft tissue injury, associated head injury, admission to ICU, and positive FAST examination can serve as reliable predictors for an elevated mortality risk in such patients.

A dual role for the transcription factor Sp8 in postnatal neurogenesis.

Neural stem cells (NSCs) of the postnatal subventricular zone (SVZ) continue producing distinct subtypes of olfactory bulb (OB) interneurons throughout life. Understanding the transcriptional coding of this diversity remains a great challenge of modern neurosciences. Interneurons expressing calretinin (CalR) represent the main interneuron subtype produced in the glomerular cell layer (GL) after birth. Previous studies have suggested that their specification relies on expression of the transcription factor Sp8 by SVZ NSCs. In this study, we performed fate mapping of NSCs that generate CalR+ or non-CalR+ interneurons, in order to assess the pattern of Sp8 expression during postnatal neurogenesis. We highlight a complex pattern of Sp8 expression, which appears to be expressed in all interneurons lineages, before getting gradually restricted to maturing CalR+ interneurons. To decipher the early and late functions of Sp8 in postnatal OB neurogenesis, we combined transient, permanent and conditional genetic approaches to manipulate Sp8 at distinct neurogenic stages. While Sp8 plays an early role in controlling proliferation in all lineages, it is not involved in the early specification of CalR+ periglomerular interneurons, but plays a crucial role in their long term survival. Together, our results highlight a crucial and dual role for Sp8 during postnatal neurogenesis.

Evaluation of the geriatric co-management for patients with fragility fractures of the proximal femur (Geriatric Fracture Centre (GFC) concept): protocol for a prospective multicentre cohort study.

INTRODUCTION: Treatment of fractures in the elderly population is a clinical challenge due partly to the presence of comorbidities. In a Geriatric Fracture Centre (GFC), patients are co-managed by a geriatrician in an attempt to improve clinical outcomes and reduce morbidity and mortality. Until now the beneficial effect of orthogeriatric co-management has not been definitively proven. The primary objective of this study is to determine the effect of GFC on predefined major adverse events related to a hip fracture compared to usual care centres (UCC). The secondary objectives include assessments in quality of life, patient-reported outcomes and cost-effectiveness. METHODS AND ANALYSIS: Two hundred and sixty-six elderly patients diagnosedwith hip fracture and planned to be treated with osteosynthesis or endoprosthesis in either a GFC or UCC study site will be recruited, 133 per type of centre. All procedures and management will be done according to the site's standard of care. Study-related visits will be performed at the following time points: preoperative, intraoperative, discharge from the orthopaedic/trauma department, discharge to definite residential status, 12 weeks and 12 months postsurgery. Data collected include demographics, residential status, adverse events, patient-reported outcomes, fall history, costs and resources related to treatment. The risk of major adverse events at 12 months will be calculated for each centre type; patient-reported outcomes will be analysed by mixed effects regression models to estimate differences in mean scores between baseline and follow-ups whereas cost-effectiveness will be assessed using the incremental cost-effectiveness ratio. ETHICS AND DISSEMINATION: Ethics approval for this study was granted from the local Ethics Committees or Institutional Review Board from each of the participating sites prior to patient enrolment. The results of this study will be published in peer-reviewed journals and presented at different conferences. TRIAL REGISTRATION NUMBER: ClinicalTrials.gov: NCT02297581; pre-results.

Improving site selection in clinical studies: a standardised, objective, multistep method and first experience results.

INTRODUCTION: A considerable number of clinical studies experience delays, which result in increased duration and costs. In multicentre studies, patient recruitment is among the leading causes of delays. Poor site selection can result in low recruitment and bad data quality. Site selection is therefore crucial for study quality and completion, but currently no specific guidelines are available. MATERIAL AND METHODS: Selection of sites adequate to participate in a prospective multicentre cohort study was performed through an open call using a newly developed objective multistep approach. The method is based on use of a network, definition of objective criteria and a systematic screening process. ILLUSTRATIVE EXAMPLE OF THE METHOD AT WORK: Out of 266 interested sites, 24 were shortlisted and finally 12 sites were selected to participate in the study. The steps in the process included an open call through a network, use of selection questionnaires tailored to the study, evaluation of responses using objective criteria and scripted telephone interviews. At each step, the number of candidate sites was quickly reduced leaving only the most promising candidates. Recruitment and quality of data went according to expectations in spite of the contracting problems faced with some sites. CONCLUSION: The results of our first experience with a standardised and objective method of site selection are encouraging. The site selection method described here can serve as a guideline for other researchers performing multicentre studies. TRIAL REGISTRATION NUMBER: ClinicalTrials.gov: NCT02297581.

Prognostic Factors in Surgical Patients with Chordomas of the Cervical Spine: A Study of 52 Cases from a Single Institution.

BACKGROUND: The recurrence rate of cervical chordomas is high, and hence it is important to discern the prognostic factors for local relapse and overall survival (OS). METHODS: A retrospective review of 52 patients with cervical chordoma operated in our institution from January 1999 to March 2015 was performed. The association of clinicopathologic characteristics with local relapse-free survival (LRFS) and OS was analyzed. Univariate analysis was performed to determine whether tumor characteristics and types of therapy affected prognosis, and a multivariate Cox proportional hazard model was developed to further investigate local recurrence and mortality. RESULTS: Mean follow-up time was 50 months. The cumulative 5- and 10-year LRFS was 35 and 0%, respectively, while the cumulative 5- and 10-year OS was 69 and 53%, respectively. The univariate analysis identified contiguous segments involved, intralesional surgical margin at primary surgery, primary surgery in local hospital, incisional biopsy, and without adjuvant radiotherapy as negative prognostic factors for LRFS, whereas for OS, only tumor location in the upper cervical spine was statistically significant. In the multivariate analysis, contiguous vertebral segments involved, intralesional surgical margins, and incisional biopsy were identified as negative prognostic factors for LRFS, whereas for OS, again only tumor location in the upper cervical spine was statistically significant. CONCLUSIONS: Contiguous vertebral segments involved, intralesional surgical margin, without adjuvant radiotherapy, and incisional biopsy significantly increase local recurrence, while tumor location in the upper cervical spine significantly increases tumor-related mortality. Thus, computed tomography-guided fine-needle aspiration biopsy and total spondylectomy with marginal excision may improve survival of patients with cervical chordoma.

Expert Consensus on Predictive Parameters for the Occurrence of a Fall and a Contralateral Hip Fracture 1 and 3 Years After Hip Fracture: A Delphi Survey.

BACKGROUND: Early identification of people at risk for a contralateral hip fracture would be desirable to favorably influence patients' prognosis. A recent systematic review failed to depict stringent patterns of risk parameters to be used for decision-making in clinical practice. OBJECTIVE: To perform a consensus study using the Delphi method to reach an expert consensus on predictive parameters for the occurrence of a fall and a contralateral hip fracture 1 and 3 years after hip fracture. METHODS: A list of potential members of the expert panel was identified based on the authors' list of a recently conducted systematic review. Participating experts were asked to name parameters determining the probability for a fall and a contralateral hip fracture 1 and 3 years after an occurred hip fracture, separately. Additionally, we asked how those stated parameters should be measured. All mentioned parameters were compiled and sent back to the experts asking them to weight each single parameter by assigning a number between 1 (not important) and 10 (very important). The survey was conducted online using the REDCap software package. We defined expert agreement if the interquartile range of attributed weights for a parameter was ≤2. A relevant parameter had at least a median weight of 8. RESULTS: Twelve experts from 7 countries completed the survey. Presence of fall history and mental and general health status were considered relevant irrespective of the outcome. For falling within 1 and 3 years, the number of medications and residential status were considered relevant, while for fractures within 1 and 3 years, osteoporosis management was considered important. CONCLUSION: Using the insights gained in this consensus study, empiric studies need to be set up assessing the prognostic value of the selected parameters.

Transcriptional Hallmarks of Heterogeneous Neural Stem Cell Niches of the Subventricular Zone.

Throughout postnatal life in mammals, neural stem cells (NSCs) are located in the subventricular zone (SVZ) of the lateral ventricles. The greatest diversity of neuronal and glial lineages they generate occurs during early postnatal life in a region-specific manner. In order to probe heterogeneity of the postnatal SVZ, we microdissected its dorsal and lateral walls at different postnatal ages and isolated NSCs and their immediate progeny based on their expression of Hes5-EGFP/Prominin1 and Ascl1-EGFP, respectively. Whole genome comparative transcriptome analysis revealed transcriptional regulators as major hallmarks that sustain postnatal SVZ regionalization. Manipulation of single genes encoding for locally enriched transcription factors (loss-of-function or ectopic gain-of-function in vivo) influenced NSC specification indicating that the fate of regionalized postnatal SVZ-NSCs can be readily modified. These findings reveal the pronounced transcriptional heterogeneity of the postnatal SVZ and provide targets to recruit region-specific lineages in regenerative contexts. Stem Cells 2015;33:2232-2242.

E-proteins orchestrate the progression of neural stem cell differentiation in the postnatal forebrain.

BACKGROUND: Neural stem cell (NSC) differentiation is a complex multistep process that persists in specific regions of the postnatal forebrain and requires tight regulation throughout life. The transcriptional control of NSC proliferation and specification involves Class II (proneural) and Class V (Id1-4) basic helix-loop-helix (bHLH) proteins. In this study, we analyzed the pattern of expression of their dimerization partners, Class I bHLH proteins (E-proteins), and explored their putative role in orchestrating postnatal subventricular zone (SVZ) neurogenesis. RESULTS: Overexpression of a dominant-negative form of the E-protein E47 (dnE47) confirmed a crucial role for bHLH transcriptional networks in postnatal neurogenesis by dramatically blocking SVZ NSC differentiation. In situ hybridization was used in combination with RT-qPCR to measure and compare the level of expression of E-protein transcripts (E2-2, E2A, and HEB) in the neonatal and adult SVZ as well as in magnetic affinity cell sorted progenitor cells and neuroblasts. Our results evidence that E-protein transcripts, in particular E2-2 and E2A, are enriched in the postnatal SVZ with expression levels increasing as cells engage towards neuronal differentiation. To investigate the role of E-proteins in orchestrating lineage progression, both in vitro and in vivo gain-of-function and loss-of-function experiments were performed for individual E-proteins. Overexpression of E2-2 and E2A promoted SVZ neurogenesis by enhancing not only radial glial cell differentiation but also cell cycle exit of their progeny. Conversely, knock-down by shRNA electroporation resulted in opposite effects. Manipulation of E-proteins and/or Ascl1 in SVZ NSC cultures indicated that those effects were Ascl1 dependent, although they could not solely be attributed to an Ascl1-induced switch from promoting cell proliferation to triggering cell cycle arrest and differentiation. CONCLUSIONS: In contrast to former concepts, suggesting ubiquitous expression and subsidiary function for E-proteins to foster postnatal neurogenesis, this work unveils E-proteins as being active players in the orchestration of postnatal SVZ neurogenesis.

Pax6 is essential for the maintenance and multi-lineage differentiation of neural stem cells, and for neuronal incorporation into the adult olfactory bulb.

The paired type homeobox 6 (Pax6) transcription factor (TF) regulates multiple aspects of neural stem cell (NSC) and neuron development in the embryonic central nervous system. However, less is known about the role of Pax6 in the maintenance and differentiation of adult NSCs and in adult neurogenesis. Using the +/Sey(Dey) mouse, we have analyzed how Pax6 heterozygosis influences the self-renewal and proliferation of adult olfactory bulb stem cells (aOBSCs). In addition, we assessed its influence on neural differentiation, neuronal incorporation, and cell death in the adult OB, both in vivo and in vitro. Our results indicate that the Pax6 mutation alters Nestin(+)-cell proliferation in vivo, as well as self-renewal, proliferation, and survival of aOBSCs in vitro although a subpopulation of +/Sey(Dey) progenitors is able to expand partially similar to wild-type progenitors. This mutation also impairs aOBSC differentiation into neurons and oligodendrocytes, whereas it increases cell death while preserving astrocyte survival and differentiation. Furthermore, Pax6 heterozygosis causes a reduction in the variety of neurochemical interneuron subtypes generated from aOBSCs in vitro and in the incorporation of newly generated neurons into the OB in vivo. Our findings support an important role of Pax6 in the maintenance of aOBSCs by regulating cell death, self-renewal, and cell fate, as well as in neuronal incorporation into the adult OB. They also suggest that deregulation of the cell cycle machinery and TF expression in aOBSCs which are deficient in Pax6 may be at the origin of the phenotypes observed in this adult NSC population.

Persistent Wnt/ß-catenin signaling determines dorsalization of the postnatal subventricular zone and neural stem cell specification into oligodendrocytes and glutamatergic neurons.

In the postnatal and adult central nervous system (CNS), the subventricular zone (SVZ) of the forebrain is the main source of neural stem cells (NSCs) that generate olfactory neurons and oligodendrocytes (OLs), the myelinating cells of the CNS. Here, we provide evidence of a primary role for canonical Wnt/ß-catenin signaling in regulating NSC fate along neuronal and oligodendroglial lineages in the postnatal SVZ. Our findings demonstrate that glutamatergic neuronal precursors (NPs) and oligodendrocyte precursors (OPs) are derived strictly from the dorsal SVZ (dSVZ) microdomain under the control of Wnt/ß-catenin, whereas GABAergic NPs are derived mainly from the lateral SVZ (lSVZ) microdomain independent of Wnt/ß-catenin. Transcript analysis of microdissected SVZ microdomains revealed that canonical Wnt/ß-catenin signaling was more pronounced in the dSVZ microdomain. This was confirmed using the ß-catenin-activated Wnt-reporter mouse and by pharmacological stimulation of Wnt/ß-catenin by infusion of the specific glycogen synthase kinase 3ß inhibitor, AR-A014418, which profoundly increased the generation of cycling cells. In vivo genetic/pharmacological stimulation or inhibition of Wnt/ß-catenin, respectively, increased and decreased the differentiation of dSVZ-NSCs into glutamatergic NPs, and had a converse effect on GABAergic NPs. Activation of Wnt/ß-catenin dramatically stimulated the generation of OPs, but its inhibition had no effect, indicating other factors act in concert with Wnt/ß-catenin to fine tune oligodendrogliogenesis in the postnatal dSVZ. These results demonstrate a role for Wnt/ß-catenin signaling within the dorsal microdomain of the postnatal SVZ, in regulating the genesis of glutamatergic neurons and OLs.

A global transcriptome analysis reveals molecular hallmarks of neural stem cell death, survival, and differentiation in response to partial FGF-2 and EGF deprivation.

Neurosphere cell culture is a commonly used model to study the properties and potential applications of neural stem cells (NSCs). However, standard protocols to culture NSCs have yet to be established, and the mechanisms underlying NSC survival and maintenance of their undifferentiated state, in response to the growth factors FGF-2 and EGF are not fully understood. Using cultures of embryonic and adult olfactory bulb stem cells (eOBSCs and aOBSCs), we analyzed the consequences of FGF-2 and EGF addition at different intervals on proliferation, cell cycle progression, cell death and differentiation, as well as on global gene expression. As opposed to cultures supplemented daily, addition of FGF-2 and EGF every 4 days significantly reduced the neurosphere volume and the total number of cells in the spheres, mainly due to increased cell death. Moreover, partial FGF-2 and EGF deprivation produced an increase in OBSC differentiation during the proliferative phase. These changes were more evident in aOBSC than eOBSC cultures. Remarkably, these effects were accompanied by a significant upregulation in the expression of endogenous Fgf-2 and genes involved in cell death and survival (Cryab), lipid catabolic processes (Pla2g7), cell adhesion (Dscaml1), cell differentiation (Dscaml1, Gpr17, S100b, Ndrg2) and signal transduction (Gpr17, Ndrg2). These findings support that a daily supply of FGF-2 and EGF is critical to maintain the viability and the undifferentiated state of NSCs in culture, and they reveal novel molecular hallmarks of NSC death, survival and the initiation of differentiation.

3-dimensional examination of the adult mouse subventricular zone reveals lineage-specific microdomains.

Recent studies suggest that the subventricular zone (SVZ) of the lateral ventricle is populated by heterogeneous populations of stem and progenitor cells that, depending on their exact location, are biased to acquire specific neuronal fates. This newly described heterogeneity of SVZ stem and progenitor cells underlines the necessity to develop methods for the accurate quantification of SVZ stem and progenitor subpopulations. In this study, we provide 3-dimensional topographical maps of slow cycling "stem" cells and progenitors based on their unique cell cycle properties. These maps revealed that both cell populations are present throughout the lateral ventricle wall as well as in discrete regions of the dorsal wall. Immunodetection of transcription factors expressed in defined progenitor populations further reveals that divergent lineages have clear regional enrichments in the rostro-caudal as well as in the dorso-ventral span of the lateral ventricle. Thus, progenitors expressing Tbr2 and Dlx2 were confined to dorsal and dorso-lateral regions of the lateral ventricle, respectively, while Mash1+ progenitors were more homogeneously distributed. All cell populations were enriched in the rostral-most region of the lateral ventricle. This diversity and uneven distribution greatly impede the accurate quantification of SVZ progenitor populations. This is illustrated by measuring the coefficient of error of estimates obtained by using increasing section sampling interval. Based on our empirical data, we provide such estimates for all progenitor populations investigated in this study. These can be used in future studies as guidelines to judge if the precision obtained with a sampling scheme is sufficient to detect statistically significant differences between experimental groups if a biological effect is present. Altogether, our study underlines the need to consider the SVZ of the lateral ventricle as a complex 3D structure and define methods to accurately assess neural stem cells or progenitor diversity and population sizes in physiological or experimental paradigms.

Lack of adrenomedullin affects growth and differentiation of adult neural stem/progenitor cells.

Adrenomedullin (AM) is a peptide hormone involved in the modulation of cellular growth, migration, apoptosis, and angiogenesis. These characteristics suggest that AM is involved in the control of neural stem/progenitor cell (NSPC) biology. To explore this hypothesis, we have obtained NSPC from the olfactory bulb of adult wild-type animals and brain conditional knockouts for adm, the gene that produces AM. Knockout NSPC contain higher levels of hyperpolymerized tubulin and more abundant filopodia than adm-containing cells, resulting in a different morphology in culture, whereas the size of the knockout neurospheres is smaller than that of the wild-types. Proliferation studies have demonstrated that adm-null NSPC incorporate less 5'-bromodeoxyuridine (BrdU) than their wild-type counterparts. In contrast, BrdU studies in the olfactory bulb of adult animals show more labeled cells in adm-null mice that in wild-types, suggesting that a compensatory mechanism exists that guarantees the sufficient production of neural cells in this organ. In NSPC differentiation tests, lack of adm results in significantly lower proportions of neurons and astrocytes and higher proportions of oligodendrocytes. The oligodendrocytes produced from adm-null neurospheres present an immature phenotype with fewer and shorter processes than adm-containing oligodendrocytes. Thus, AM is an important factor in regulating the proliferation and differentiation of adult NSPC and might be used to modulate stem cell renewal and fate in protocols destined to produce neural cells for regenerative therapies.

IGF-I promotes neuronal migration and positioning in the olfactory bulb and the exit of neuroblasts from the subventricular zone.

While insulin-like growth factor-I (IGF-I) supports neuronal and glial differentiation in the CNS, it is largely unknown whether IGF-I also influences neuronal migration and positioning. We show here that the pattern of olfactory bulb (OB) layering is altered in Igf-I (-/-) mice. In these animals, Tbr1(+)-glutamatergic neurons are misplaced in the mitral cell layer (ML) and the external plexiform layer (EPL). In addition, there are fewer interneurons in the glomerular layer and the EPL of the Igf-I (-/-) mice, and fewer newborn neurons are incorporated into the OB from the forebrain subventricular zone (SVZ). Indeed, neuroblasts accumulate in the postnatal/adult SVZ of Igf-I (-/-) mice. Significantly, the positioning of Tbr1(+)-cells in a primitive ML is stimulated by IGF-I in cultured embryonic OB slices, an effect that is partially repressed by the phosphoinositide 3-kinase (PI3K) inhibitor. In OB cell cultures, IGF-I increases the phosphorylation of disabled1 (P-Dab1), an adaptor protein that is a target of Src family kinases (SFK) in the reelin signalling pathway, whereas reduced P-Dab1 levels were found in Igf-I (-/-) mice. Neuroblast migration from the rostral migratory stream (RMS) explants of postnatal Igf-I (-/-) was similar to that from Igf-I (+/+) explants. However, cell migration was significantly enhanced by IGF-I added to the explants, an effect that was repressed by PI3K and SFK inhibitors. These findings suggest that IGF-I promotes neuronal positioning in the OB and support a role for IGF-I in stimulating neuroblast exit from the SVZ into the RMS, thereby promoting the incorporation of newly formed neurons into the OB.