scispace - formally typeset

Posted ContentDOI

In - Vivo Conversion of Astrocytes to Neuroblasts in the Injured Spinal Cord

26 Apr 2021-

TL;DR: Maryam Lale Ataei Tabriz Medical University: Tabriz University of Medical Sciences Mohammad Karimipour (  karimiportm@tbzmed.ac.ir) and Parviz Shahabi are the authors of this monograph.
Abstract: Maryam Lale Ataei Tabriz Medical University: Tabriz University of Medical Sciences Mohammad Karimipour (  karimipourm@tbzmed.ac.ir ) Tabriz University of Medical Sciences https://orcid.org/0000-0002-6975-4308 Parviz Shahabi Tabriz Medical University: Tabriz University of Medical Sciences Hamid Soltani-Zangbar Tabriz Medical University: Tabriz University of Medical Sciences Maryam Pashaiasl Tabriz Medical University: Tabriz University of Medical Sciences
Topics: Spinal cord (54%), Neuroblast (54%)

Summary (4 min read)

Introduction

  • Spinal cord injury (SCI) is damage to the spinal cord and a complex neurological condition that resulted in irreversible neuronal loss, glial scar formation, axonal injury, disruption of the myelin sheath and neural tracts and long-lasting disability and in some cases leads to permanent functional de cits in mammals (Su et al.
  • It is shown that the SRY (sex-determining region Y-box 2), also known as Sox2, is a transcription factor that is essential for maintaining self-renewal, or pluripotency, of undifferentiated embryonic stem cells.
  • Sox2 has a critical role in the maintenance of embryonic and neural stem cells and also it is crucial for directing neural differentiation (Zhang and Cui 2014; Mercurio et al.
  • Astrocytes due to nearness to radial glial cells identity, their high quantity, and the potential to proliferate in the central nervous system, astrocytes could undertake therapeutic interventional approaches including reprogramming.
  • The concept of direct reprogramming is a process in which one mature somatic cell transforms into another mature somatic cell without undergoing an intermediate pluripotent state or progenitor cell type(Ghasemi- Kasman et al. 2015) .

Ethical issue and study design

  • In the current study, 54 adult male Wister rats (weight: 270-300g) were purchased from the animal laboratory and maintained according to the guide line of ethics committee of Tabriz University of Medical Sciences (registered number 95/5-10/7).
  • All animals were housed in a standard condition under a 12h light/dark schedule with enough food and water.
  • In all groups, laminectomy was performed at the T9-T10 vertebral level in the dorsal surface of the spinal cord using the In nite Horizons Impactor with an impact force of 150 (moderate SCI) kdyn (1 dyn=1g⋅cm/s 2=10-5kg⋅m/s 2=10-5 N) by impactor device.
  • Amniotic uid extraction carried out under supervision the gynecologist with using a 22G Needle.
  • In the cultivation period, the medium was changed twice a week, and in the 90% con uence, the cells trypsinized with trypsin-EDTA [0.25%] and centrifuged, and nally, cells pellet re-seeded in DMEM-low glucose media with 15% FBS, 1% penicillin/streptomycin and10 ng/mL bFGF in the optimized condition.

Preparation of CM

  • The MSCs at the 3rd-5rd passage and 90% con uent, were trypsinized and washed with PBS three times, and re-fed with DMEM-low glucose culture medium in serum-free condition for 48 h.
  • Finally, CM was sterilized through 0.22 µm lters and concentrated by freeze-drying processes and was stored at -80 ℃ until use.

Western blot

  • Sox2 (Sex determining region Y-box 2) secreted by HAF-MSCs into MSCs-CM, was measured using western blot analysis.

Human astrocyte culture

  • Human astrocytes (line 1321N1) (Ghasemi-Kasman et al. 2015) were purchased from Pasteur Institute of Iran and cultured in DMEM low glucose medium with 10% FBS and 1% penicillin/streptomycin.

Spinal Cord Surgery

  • After deep anesthesia, an adequate level of anesthesia was determined by checking withdrawal to painful stimuli applied to the hind limb.
  • The rats were shifted to the surgical location and via the mask received an iso urane vapor inhalation (3-5%) and oxygen (0.8-1 L/min) to the end of surgical procedure.
  • Brie y, animals under anesthesia conditions, their back was shaved and in the midline, the skin s incision was performed around 2cm, and in order to laminectomy, Paravertebral muscles were cut up on the T9-T11 spinal processes, and with the dental drill, a hole of 1.5 mm diameter was made in the vertebral arch of T10 as far as dura mater could be seen.
  • To prevent infection following SCI, cipro oxacin (350 ml units/days) was injected via IP for one week.
  • After SCI, the bladder sac was discharged manually twice a day for one week.

Infusion of MSCs-CM

  • In order to explore the effect of the CM on the rate expression of the endogenous neuroblasts and astrocytes, 500 µl of CM, following SCI was infused through intraperitoneal (IP) per day for 7 days.

Transplantation of hAF-MSCs

  • The next, to examine the effect of the hAF-MSCs on endogenous expression of neuroblasts and astrocytes, the MSCs were detached and harvested using 0.25% Trypsin-EDTA.
  • Prior to the transplantation, the number of cells was estimated by counting in a neobar lam and washed by PBS three times.
  • Following SCI, 5×105 cells per 5µl PBS were transplanted to the proximal, central, and distal parts of the lesion site using a capillary glass needle through a Hamilton syringe.
  • For immunosuppression, the rats received cyclosporine (1 mg/100 g body weight) two days before cell transplantation until the end of the experiment (Springer et al. 2018 ).

Injection of the exogenous human astrocytes

  • In the next series of experiments, the authors decided to investigate the effect of the MSCs and their CM on astrocyte reprogramming and converting the human astrocytes to neuroblasts.
  • To this end, the human astrocytes (Cell line 1321N1) were injected focally into the lesion site of the spinal cord concomitantly with transplantation of MSCs and infusion of CM.

Tissue preparation and immuno uorescence staining

  • For immuno uorescence examinations, animals were sacri ced two weeks after SCI by ketamine (100mg/kg) and xylazine (5mg/kg) overdose.
  • The rats were transcardially perfused with normal saline (NaCl 9%) and 4% paraformaldehyde, respectively.
  • The animal's spinal cord was carefully removed and post-xed overnight with 4% paraformaldehyde solution.
  • After three times PBS wash, the sections were incubated with secondary antibodies including Goat Anti-Mouse IgG H&L (ab 97024), Goat Anti-Rabbit IgG H&L (FITC) (ab 6717), Donkey Anti-Goat IgG H&L (FITC) (ab6881) and Goat Anti-Mouse IgG H&L (Texas Red ®) (ab6787) at room temperature for 1 hour.
  • Also, nuclei were counterstained with 4′, 6diamidino-2-phenylindole (DAPI) (ab 104139).

Neurobehavioral examination

  • Locomotor performance on 1, 7 and 21 days after SCI was assessed with the use of the 21 point (a score from 0 (complete paralysis) to 21 (normal gait)) BBB (Basso, Beattie and Bresnahan) score by two examiners blinded.

Statistical analysis

  • One-way analysis of variance and post hoc.
  • Tukey tests were performed to detect the statistically signi cant difference between groups.
  • P<0.05 was considered as statistically signi cant.
  • All the statistics presented in the article were analyzed and drawn using Graph Pad Prism (version 6.01; Graph Pad Software, CA, USA).

Flow cytometry analysis and phenotype acquisition

  • Isolated and cultivated of MSCs from human amniotic uid showed spindle-like morphology and broblast-like under bright-eld microscope.
  • Evaluation of cell surface markers of the MSCs at Passages (3-5) by ow cytometry analysis revealed that the MSCs express CD105 (95.80%) and CD73 (92.2%) as the MSCs markers while they didn't express, CD14 (9.90%) and CD45 (9.16%) as the hematopoietic markers .

MSCs secreted the SOX2 protein in the CM

  • This analysis showed that the Sox2 protein was observed in the 32 folds concentrated CM.
  • The results from this panel showed that the MSCs after two weeks of transplantation promoted the level of DCX expression and suppressed the rate of GFAP marker.
  • In general, these data indicate the MSCs through juxtacrine and paracrine pathways could promote the neuroblasts and induce neurogenesis, as well as, diminish the astrocytes in the SCI and nally suppress gliosis and glial scar formation.
  • MSCs and CM could increase the number of neuroblasts relative to the (SCI +Astrocytes + DMEM) group (p<.001, p<.05) respectively .
  • Moreover, presented data showed that MSCs in comparison to CM could raise the functional recovery score more than the CM(p<.05) .

Discussion

  • Astrocytes in large quantities have been distributed throughout the CNS and provide essential factors and desire microenvironment for optimal neural tissue structure and function in a healthy condition (Sofroniew and Vinters 2010) .
  • Regardless of these ndings, astrocyte-derived neurons have not identi ed around the injured area in both the brain and spinal cord (Ohori et al.
  • Therefore, nding new and effective therapeutic strategies including reprogramming and neurogenesis promotion is a challenging and debate subject in the regenerative medicine eld.
  • According to previous evidence, transcription factor Sox2 could be su cient to converting endogenous differentiated astrocytes into neuroblasts and also mature neurons in the adult spinal cord with different severity of the injury (Su et al. 2014a ).
  • In conclusion, the results of the present research point out the protective and regenerative potential of MSCs in the SCI through reprogramming astrocytes to neuroblasts by mediating juxtacrine activity and paracrine effects.

Declarations

  • Author Contributions M.PA and P.SH designed and supervised the whole study.
  • M.K prepared and wrote the manuscript and analyzed data.
  • M.A performed the experiments and involved in data acquisition.

Funding:

  • This work was supported by a grant (No: 95/5-10/7) from Tabriz University of Medical Sciences.

Compliance with Ethical

  • The authors declare that they have no competing interests.

Ethical Approval

  • All experiments were conducted according to international principal guidelines and approved by a local ethics committee of Tabriz University of Medical Sciences.

Did you find this useful? Give us your feedback

Content maybe subject to copyright    Report

Page 1/17
In - Vivo
Conversion of Astrocytes to Neuroblasts in
the Injured Spinal Cord
Maryam Lale Ataei
Tabriz Medical University: Tabriz University of Medical Sciences
Mohammad Karimipour ( karimipourm@tbzmed.ac.ir )
Tabriz University of Medical Sciences https://orcid.org/0000-0002-6975-4308
Parviz Shahabi
Tabriz Medical University: Tabriz University of Medical Sciences
Hamid Soltani-Zangbar
Tabriz Medical University: Tabriz University of Medical Sciences
Maryam Pashaiasl
Tabriz Medical University: Tabriz University of Medical Sciences
Research Article
Keywords: Spinal cord injury, MSCs, Astrocyte, Neuroblast, Reprogramming
Posted Date: April 26th, 2021
DOI: https://doi.org/10.21203/rs.3.rs-362556/v1
License: This work is licensed under a Creative Commons Attribution 4.0 International License.
Read Full License

Page 2/17
Abstract
Direct astrocyte reprogramming to neural progenitor cells and promotion of neurogenesis is considered
as an alternative approach to replace the lost neurons in the spinal cord injury(SCI). Herein, we used the
human amniotic uid mesenchymal stem cells (hAF-MSCs) and their conditioned medium (CM), to
investigating their ability to reprogramming astrocytes to neuroblasts following SCI. 54 adult rats were
randomly divided into 9 groups (n = 6), included: Control, SCI, (SCI + DMEM), (SCI + CM), (SCI+ MSCs),
(SCI + Astrocyte), (SCI + Astrocyte + DMEM), (SCI+ Astrocyte+ CM) and SCI+ Astrocyte+ MSCs).
Following laminectomy and SCI induction, DMEM, CM, MSCs and Astrocytes were injected. Wester-blot
was performed to explore the levels of the Sox2 protein in the MSCs-CM. The immunouorescence
staining against DCX and GFAP was done. Finally, Basso-Beattie-Brenham (BBB) locomotor test was
conducted to assess the neurological outcomes. Our results showed that the MSCs through juxtacrine
and paracrine mechanisms induced the promotion of the endogenous neuroblasts and the decline of
astrocytes. Moreover, in the present research, MSCs and CM could convert the transplanted human
astrocytes to neuroblasts in the spinal cord injury. Taken together, our data indicate the MSCs via
juxtacrine and paracrine pathways could direct the spinal cord endogenous neural stem cells(NSCs) to
the neuroblasts lineage rather than astrocytes as well as induce reprogramming. Ultimately, MSCs could
reverse the neurobehavioral decit in the SCI. The striking output in our study was the capability of the
MSCs in the reprogramming of the astrocytes to neuroblasts via juxtacrine and paracrine pathways in the
In-vivo
condition.
Introduction
Spinal cord injury (SCI) is damage to the spinal cord and a complex neurological condition that resulted
in irreversible neuronal loss, glial scar formation, axonal injury, disruption of the myelin sheath and neural
tracts and long-lasting disability and in some cases leads to permanent functional decits in
mammals(Su et al. 2014b; Higuchi et al. 2019; Abolhasanpour et al. 2019). Following SCI, astrocytes
proliferate and migrate into the surrounding milieu and generate glial scar and undesirable
microenvironment (Wang et al. 2016). The research demonstrated that the astrocytes and NG2 (Pre-
oligodrocytes), as a result of reprogramming could convert to neuroblast and neurons in the spinal cord
and brain (Wang et al. 2016). It is shown that the SRY (sex- determining region Y-box 2), also known as
Sox2, is a transcription factor that is essential for maintaining self-renewal, or pluripotency, of
undifferentiated embryonic stem cells. Sox2 has a critical role in the maintenance of embryonic and
neural stem cells and also it is crucial for directing neural differentiation (Zhang and Cui 2014; Mercurio
et al. 2019; Rodriguez-Jimenez et al. 2016). Sox2 can mediated reprogramming of astrocytes to DCX +
neuroblasts and in this context, Sox2 can help to treat SCI by converting glial cells to neurons nally
mature neurons (Wang et al. 2016; Rodriguez-Jimenez et al. 2016). Doublecortin (DCX) is a microtubule-
associated protein expressed by neuronal precursor cells and immature neurons in embryonic and adult
cortical structures and considered as a marker of developing neural progenitor cells (Karimipour et al.
2019; Nasrolahi et al. 2019). Astrocytes due to nearness to radial glial cells identity, their high quantity,

Page 3/17
and the potential to proliferate in the central nervous system, astrocytes could undertake therapeutic
interventional approaches including reprogramming. (Sun et al. 2019). Recent advancements in the eld
of direct in vivo reprogramming showed the generation of functional neurons from reactive glial cells in
the repair of the brain (Li and Chen 2016; Guo et al. 2014). The concept of direct reprogramming is a
process in which one mature somatic cell transforms into another mature somatic cell without
undergoing an intermediate pluripotent state or progenitor cell type(Ghasemi-Kasman et al. 2015).
Transplantation stem cells including MSCs and olfactory ensheathing cells (OECs) in SCI could lead to
increase of neuroprotective molecules and reprogramming of astrocytes into neuroblasts which could
survive and develop into mature neurons (Sabapathy et al. 2017; Sun et al. 2019). In this context, MSCs
by secretion of the essential materials for neuroprotection such as growth factors, cytokines, and
neurotrophic factors, could induce the replacement of lost neuronal cells, remyelination of axons,
angiogenesis, and decline of inammatory responses(Oh and Jeon 2016; Shahrezaie et al. 2017). MSCs
could harvest from different sources including adipose derived tissue, bone marrow, Umbilical Cord
Wharton's Jelly, amniotic membrane and amniotic uid mesenchymal stem cells (hAF-MSCs) and so on
(Gabel et al. 2017; Oh and Jeon 2016). In spite of the plethora of studies and experiments regarding the
evaluation of the functional ecacy of the MSCs in the SCI, the election of the effective source of the
MSCs is one of the challenging issues in the scientic community (Gabel et al. 2017). But recent studies,
suggest that (hAF-MSCs) could account into consideration in the SCI because of the amnion origin from
the epiblast layer as a pluripotent and undifferentiated structure in the embryo(De Coppi et al. 2007;
Antonucci et al. 2014). These cells routinely gained during amniocentesis which is done in the second
trimester and express embryonic and pluripotency markers for instance Oct4, Nanog and Sox2 (Aziz et al.
2019; Gholizadeh-Ghaleh Aziz et al. 2019; Dziadosz et al. 2016; Maraldi et al. 2014; Saito et al. 2012).
Amniotic uid derived- MSCs relative to other sources have high proliferative and regenerative potential
as well as they maintain pluripotency features, and remain without alteration after continuous passages
(Maraldi et al. 2014; Aziz et al. 2016). MSCs during cell culture can secrete paracrine factors in the form
of conditioned medium (CM) (Osugi et al. 2012; Cantinieaux et al. 2013). CM contains metabolites,
growth factors, extracellular matrix proteins, cytokines and anti-inammatory agents which are secreted
into the medium by the cultured cells. (Cantinieaux et al. 2013; Pawitan 2014). Sox2 which is available in
hAF-MSCs-CM is the essential factor for the transform endogenous spinal astrocytes to neuroblasts (Su
et al. 2014b; Zhang and Cui 2014). In this Perspective, we investigated the eciency of the hAF-MSCs
and their CM in the reprogramming of the astrocytes to neuroblasts through juxtacrine and paracrine
mechanisms as well as functional behavior following the SCI.
Materials And Methods
Ethical issue and study design
In the current study, 54 adult male Wister rats (weight: 270-300g) were purchased from the animal
laboratory and maintained according to the guide line of ethics committee of Tabriz University of Medical
Sciences (registered number 95/5-10/7). All animals were housed in a standard condition under a 12h
light/dark schedule with enough food and water. The rats were randomly divided in to 9 groups (six rats

Page 4/17
per group), these nine groups included:Control,SCI, (SCI + DMEM, IP), (SCI + CM, IP), (SCI + MSCs,
Focally), (SCI + Astrocyte), (SCI + Astrocyte+ DMEM, IP), (SCI + Astrocyte+ CM, IP) and SCI + Astrocyte+
MSCs). In all groups, laminectomy was performed at the T9–T10 vertebral level in the dorsal surface of
the spinal cord using the Innite Horizons Impactor with an impact force of 150 (moderate SCI) kdyn (1
dyn=1gcm/s 2=10-5kgm/s 2=10-5 N) by impactor device.
hAF-MSCs isolation, cultivation and characterization
Briey, isolation of hAF-MSCs was done undergoing amniocentesis for the routine karyotype screening of
about 5 ml of amniotic uid samples from eight mothers in Al-Zahra hospital (Tabriz, Iran). Prior to the
amniocentesis, patients written informed consent for donating amniotic uid samples voluntarily for this
research. Amniotic uid extraction carried out under supervision the gynecologist with using a 22G
Needle. After sending the samples to the desired laboratory, samples were centrifuged at 450 g for 10
minutes, next the settled pellet was washed twice by PBS (Gibco; Thermo Fisher Scientic, Darmstadt,
Germany). Then, the cells transferred in 6 well plates with AmnioMAX II Complete Medium (Gibco, cat#
11269) for 1-2 weeks in condition 37°C and 5% CO2. In the cultivation period, the medium was changed
twice a week, and in the 90% conuence, the cells trypsinized with trypsin-EDTA [0.25%] (Gibco) and
centrifuged, and nally, cells pellet re-seeded in DMEM-low glucose media with 15% FBS, 1%
penicillin/streptomycin and10 ng/mL bFGF in the optimized condition. The phenotype prole of the hAF-
MSCs was examined by ow cytometry analysis. For this purpose, the cells at passage 3-5 were
trypsinzed and two times washed with PBS and centrifuge at 1500 RPM for 3 min, then, the cell pellets
were stained with antibodies including CD 105 (Catalog No. 1p-298-To25 Exbio), CD 73 (Catalog No.
561260 BD Biosciences) antibodies as mesenchymal stem cell markers and CD 45 (Catalog No. 1F-222-
T025 Exbio), CD 14 (Catalog No. 12-0149-42 eBioscience) as hematopoietic stem cell markers) with
dilution 1/30 in the PBS for 20 min on ice.
Preparation of CM
The MSCs at the 3rd-5rd passage and 90% conuent, were trypsinized and washed with PBS three times,
and refed with DMEM-low glucose culture medium in serumfree condition for 48 h. Then, CM was
harvested and centrifuged at 450g for 10 min up to eliminate free-oating cells. Finally, CM was sterilized
through 0.22 µm lters and concentrated by freeze-drying processes and was stored at -80 until use.
Western blot
CM was collected from MSCs culture and was sterilized by 0.22 mm lters, then CM was concentrated 2,
4, 8, 16 and 32 folds by freeze dryer device. Sox2 (Sex determining region Y-box 2) secreted by HAF-MSCs
into MSCs-CM, was measured using western blot analysis.
Human astrocyte culture
Human astrocytes (line 1321N1) (Ghasemi-Kasman et al. 2015) were purchased from Pasteur Institute of
Iran and cultured in DMEM low glucose medium with 10% FBS and 1% penicillin/streptomycin. This

Page 5/17
medium was changed twice in a week.
Spinal Cord Surgery
The rats were anesthetized by inhalation of 5% isourane and oxygen (1 L/min) in a closed space. After
deep anesthesia, an adequate level of anesthesia was determined by checking withdrawal to painful
stimuli applied to the hind limb. The rats were shifted to the surgical location and via the mask received
an isourane vapor inhalation (3-5%) and oxygen (0.8-1 L/min) to the end of surgical procedure. Briey,
animals under anesthesia conditions, their back was shaved and in the midline, the skin s incision was
performed around 2cm, and in order to laminectomy, Paravertebral muscles were cut up on the T9-T11
spinal processes, and with the dental drill, a hole of 1.5 mm diameter was made in the vertebral arch of
T10 as far as dura mater could be seen. Then, using the Horizons Impactor, animals received a force of
150-kilodyne (moderate SCI) on the targeted spinal cord segments, subsequently, the muscles and skin
were closed. Also for bilateral injury, transverse process of the rats throughout the surgery and injury xed
by a clamp. To prevent infection following SCI, ciprooxacin (350 ml units/days) was injected via IP for
one week. After SCI, the bladder sac was discharged manually twice a day for one week.
Infusion of MSCs- CM
In order to explore the effect of the CM on the rate expression of the endogenous neuroblasts and
astrocytes, 500 µl of CM, following SCI was infused through intraperitoneal (IP) per day for 7 days.
Transplantation of hAF-MSCs
The next, to examine the effect of the hAF-MSCs on endogenous expression of neuroblasts and
astrocytes, the MSCs were detached and harvested using 0.25% Trypsin-EDTA. Prior to the
transplantation, the number of cells was estimated by counting in a neobar lam and washed by PBS three
times. Following SCI, 5×105 cells per 5µl PBS were transplanted to the proximal, central, and distal parts
of the lesion site using a capillary glass needle through a Hamilton syringe. For immunosuppression, the
rats received cyclosporine (1 mg/100 g body weight) two days before cell transplantation until the end of
the experiment(Springer et al. 2018).
Injection of the exogenous human astrocytes
In the next series of experiments, we decided to investigate the effect of the MSCs and their CM on
astrocyte reprogramming and converting the human astrocytes to neuroblasts. To this end, the human
astrocytes (Cell line 1321N1) were injected focally into the lesion site of the spinal cord concomitantly
with transplantation of MSCs and infusion of CM.
Tissue preparation and immunouorescence staining
For immunouorescence examinations, animals were sacriced two weeks after SCI by ketamine
(100mg/kg) and xylazine (5mg/kg) overdose. The rats were transcardially perfused with normal saline

References
More filters

Journal ArticleDOI
01 Jan 2006-Cytotherapy
TL;DR: The Mesenchymal and Tissue Stem Cell Committee of the International Society for Cellular Therapy proposes minimal criteria to define human MSC, believing this minimal set of standard criteria will foster a more uniform characterization of MSC and facilitate the exchange of data among investigators.
Abstract: The considerable therapeutic potential of human multipotent mesenchymal stromal cells (MSC) has generated markedly increasing interest in a wide variety of biomedical disciplines. However, investig...

12,805 citations


"In - Vivo Conversion of Astrocytes ..." refers background or result in this paper

  • ...Overall, this experiment revealed that the MSCs and CM could increase the DCX – positive cells and diminish the GFAP-positive cells in the in-vivo condition and MSCs Page 11/17 and CM, could be considered as future relevant tools in the eld of regenerative medicine (Dominici et al. 2006)....

    [...]

  • ...The expression of the cell surface markers of the Page 10/17 isolated and cultivated MSCs in the present study was consistent with the previous researches (Markmee et al. 2017; Dominici et al. 2006), So that, the MSCs at 3–4 passage, were positive for MSCs including CD105 (95.80...

    [...]


Journal ArticleDOI
TL;DR: Astrocyte functions in healthy CNS, mechanisms and functions of reactive astrogliosis and glial scar formation, and ways in which reactive astrocytes may cause or contribute to specific CNS disorders and lesions are reviewed.
Abstract: Astrocytes are specialized glial cells that outnumber neurons by over fivefold. They contiguously tile the entire central nervous system (CNS) and exert many essential complex functions in the healthy CNS. Astrocytes respond to all forms of CNS insults through a process referred to as reactive astrogliosis, which has become a pathological hallmark of CNS structural lesions. Substantial progress has been made recently in determining functions and mechanisms of reactive astrogliosis and in identifying roles of astrocytes in CNS disorders and pathologies. A vast molecular arsenal at the disposal of reactive astrocytes is being defined. Transgenic mouse models are dissecting specific aspects of reactive astrocytosis and glial scar formation in vivo. Astrocyte involvement in specific clinicopathological entities is being defined. It is now clear that reactive astrogliosis is not a simple all-or-none phenomenon but is a finely gradated continuum of changes that occur in context-dependent manners regulated by specific signaling events. These changes range from reversible alterations in gene expression and cell hypertrophy with preservation of cellular domains and tissue structure, to long-lasting scar formation with rearrangement of tissue structure. Increasing evidence points towards the potential of reactive astrogliosis to play either primary or contributing roles in CNS disorders via loss of normal astrocyte functions or gain of abnormal effects. This article reviews (1) astrocyte functions in healthy CNS, (2) mechanisms and functions of reactive astrogliosis and glial scar formation, and (3) ways in which reactive astrocytes may cause or contribute to specific CNS disorders and lesions.

3,422 citations


"In - Vivo Conversion of Astrocytes ..." refers background in this paper

  • ...…and nally could rescue the functional de cit in the SCI. Discussion Astrocytes in large quantities have been distributed throughout the CNS and provide essential factors and desire microenvironment for optimal neural tissue structure and function in a healthy condition(Sofroniew and Vinters 2010)....

    [...]


Journal ArticleDOI
Michael V. Sofroniew1Institutions (1)
TL;DR: Developments in the signaling mechanisms that regulate specific aspects of reactive astrogliosis are reviewed and the potential to identify novel therapeutic molecular targets for diverse neurological disorders is highlighted.
Abstract: Reactive astrogliosis, whereby astrocytes undergo varying molecular and morphological changes, is a ubiquitous but poorly understood hallmark of all central nervous system pathologies. Genetic tools are now enabling the molecular dissection of the functions and mechanisms of reactive astrogliosis in vivo. Recent studies provide compelling evidence that reactive astrogliosis can exert both beneficial and detrimental effects in a context-dependent manner determined by specific molecular signaling cascades. Reactive astrocytes can have both loss of normal functions and gain of abnormal effects that could feature prominently in a variety of disease processes. This article reviews developments in the signaling mechanisms that regulate specific aspects of reactive astrogliosis and highlights the potential to identify novel therapeutic molecular targets for diverse neurological disorders.

1,948 citations


"In - Vivo Conversion of Astrocytes ..." refers background in this paper

  • ...In response to any CNS damage, they are activated, proliferated, migrated to the lesion site, and participate in glial scar formation (Sofroniew 2009; Yiu and He 2006; Buffo et al. 2008)....

    [...]


Journal ArticleDOI
Paolo De Coppi1, Georg Bartsch1, M. Minhaj Siddiqui1, Tao Xu1  +9 moreInstitutions (2)
TL;DR: The isolation of human and rodent amniotic fluid–derived stem (AFS) cells that express embryonic and adult stem cell markers are reported and examples of differentiated cells derived from human AFS cells and displaying specialized functions include neuronal lineage cells secreting the neurotransmitter L-glutamate or expressing G-protein-gated inwardly rectifying potassium channels.
Abstract: Stem cells capable of differentiating to multiple lineages may be valuable for therapy. We report the isolation of human and rodent amniotic fluid-derived stem (AFS) cells that express embryonic and adult stem cell markers. Undifferentiated AFS cells expand extensively without feeders, double in 36 h and are not tumorigenic. Lines maintained for over 250 population doublings retained long telomeres and a normal karyotype. AFS cells are broadly multipotent. Clonal human lines verified by retroviral marking were induced to differentiate into cell types representing each embryonic germ layer, including cells of adipogenic, osteogenic, myogenic, endothelial, neuronal and hepatic lineages. Examples of differentiated cells derived from human AFS cells and displaying specialized functions include neuronal lineage cells secreting the neurotransmitter L-glutamate or expressing G-protein-gated inwardly rectifying potassium channels, hepatic lineage cells producing urea, and osteogenic lineage cells forming tissue-engineered bone.

1,765 citations


Journal ArticleDOI
TL;DR: The transient expression pattern of DCX in neuronal committed progenitor cells/neuroblasts indicates that DCX could be developed into a suitable marker for adult neurogenesis and may provide an alternative to BrdU labeling.
Abstract: During development of the central nervous system, expression of the microtubule binding protein doublecortin (DCX) is associated with migration of neuroblasts. In addition to this developmental role, expression of DCX remains high within certain areas of the adult mammalian brain. These areas, mainly the dentate gyrus and the lateral ventricle wall in conjunction with the rostral migratory stream and olfactory bulb, retain the capacity to generate new neurons into adulthood. Adult neurogenesis is typically detected by incorporation of bromodeoxyuridine (BrdU) into dividing cells and colabeling of BrdU-positive cells with markers for mature neurons. To elucidate whether DCX could act as an alternative indicator for adult neurogenesis, we investigated the temporal expression pattern of DCX in neurogenic regions of the adult brain. Analysis of newly generated cells showed that DCX is transiently expressed in proliferating progenitor cells and newly generated neuroblasts. As the newly generated cells began expressing mature neuronal markers, DCX immunoreactivity decreased sharply below the level of detection and remained undetectable thereafter. The transient expression pattern of DCX in neuronal committed progenitor cells/neuroblasts indicates that DCX could be developed into a suitable marker for adult neurogenesis and may provide an alternative to BrdU labeling. This assumption is further supported by our observation that the number of DCX-expressing cells in the dentate gyrus was decreased with age according to the reduction of neurogenesis in the aging dentate gyrus previously reported.

1,398 citations


"In - Vivo Conversion of Astrocytes ..." refers background in this paper

  • ...DCX is a microtubule-associated protein expressed predominantly in neuroblasts, immature neurons in developmental processes, and most importantly, in the neurogenic area of the adult brain (Brown et al. 2003; Gleeson et al. 1999)....

    [...]